sybr green i  (Thermo Fisher)


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    Structured Review

    Thermo Fisher sybr green i
    Visual detection of amplified LAMP products using <t>SYBR</t> green I. Addition of 1 µl of diluted SYBR <t>green</t> I to the reaction tube after LAMP reaction enables visible analysis of the results under natural light ( Figure 1A ) or UV irradiation ( Figure 1B ). The color changes from orange (negative reaction) to green (positive reaction) ( Figure 1A ) and bright fluorescence indicates a positive reaction ( Figure 1B ).
    Sybr Green I, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 803 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/sybr green i/product/Thermo Fisher
    Average 99 stars, based on 803 article reviews
    Price from $9.99 to $1999.99
    sybr green i - by Bioz Stars, 2020-02
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    Images

    1) Product Images from "Evaluation of a Direct Reverse Transcription Loop-Mediated Isothermal Amplification Method without RNA Extraction for the Detection of Human Enterovirus 71 Subgenotype C4 in Nasopharyngeal Swab Specimens"

    Article Title: Evaluation of a Direct Reverse Transcription Loop-Mediated Isothermal Amplification Method without RNA Extraction for the Detection of Human Enterovirus 71 Subgenotype C4 in Nasopharyngeal Swab Specimens

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0052486

    Visual detection of amplified LAMP products using SYBR green I. Addition of 1 µl of diluted SYBR green I to the reaction tube after LAMP reaction enables visible analysis of the results under natural light ( Figure 1A ) or UV irradiation ( Figure 1B ). The color changes from orange (negative reaction) to green (positive reaction) ( Figure 1A ) and bright fluorescence indicates a positive reaction ( Figure 1B ).
    Figure Legend Snippet: Visual detection of amplified LAMP products using SYBR green I. Addition of 1 µl of diluted SYBR green I to the reaction tube after LAMP reaction enables visible analysis of the results under natural light ( Figure 1A ) or UV irradiation ( Figure 1B ). The color changes from orange (negative reaction) to green (positive reaction) ( Figure 1A ) and bright fluorescence indicates a positive reaction ( Figure 1B ).

    Techniques Used: Amplification, SYBR Green Assay, Irradiation, Fluorescence

    2) Product Images from "Rapid Detection of Zika Virus in Urine Samples and Infected Mosquitos by Reverse Transcription-Loop-Mediated Isothermal Amplification"

    Article Title: Rapid Detection of Zika Virus in Urine Samples and Infected Mosquitos by Reverse Transcription-Loop-Mediated Isothermal Amplification

    Journal: Scientific Reports

    doi: 10.1038/s41598-018-22102-5

    ZIKV and 18S rRNA RT-LAMP in simulated urine samples. Urine samples were spiked with either different strains of ZIKV or DENV and subjected to a ZIKV ( A ) or 18S rRNA ( B ) specific RT-LAMP reaction then visualized by addition of SYBR Green I by eye (upper panel), green fluorescence (middle panel), or gel electrophoresis (bottom panel). Lane M: Low DNA Mass Marker (ThermoFisher Scientific); NTC: No template control (negative control). Relative density (RD) of the entire bands for a column relative to the DNA Mass Marker are indicated below the corresponding lane. Lanes that did not have any detectable bands over background are reported as not detectable (ND).
    Figure Legend Snippet: ZIKV and 18S rRNA RT-LAMP in simulated urine samples. Urine samples were spiked with either different strains of ZIKV or DENV and subjected to a ZIKV ( A ) or 18S rRNA ( B ) specific RT-LAMP reaction then visualized by addition of SYBR Green I by eye (upper panel), green fluorescence (middle panel), or gel electrophoresis (bottom panel). Lane M: Low DNA Mass Marker (ThermoFisher Scientific); NTC: No template control (negative control). Relative density (RD) of the entire bands for a column relative to the DNA Mass Marker are indicated below the corresponding lane. Lanes that did not have any detectable bands over background are reported as not detectable (ND).

    Techniques Used: SYBR Green Assay, Fluorescence, Nucleic Acid Electrophoresis, Marker, Negative Control

    ZIKV RT-LAMP sensitivity for ZIKV. Sensitivity assessment of ZIKV RT-LAMP using serial dilutions of ZIKV PCR Standard (Robert Koch Institute) from 4 × 10 5 genome copies/reaction to 1 copies/reaction as visualized by addition of SYBR Green I by eye (upper panel), green fluorescence (middle panel), or gel electrophoresis (bottom panel). Lane M: Low DNA Mass Marker (ThermoFisher Scientific); NTC: No template control (negative control). Relative density (RD) of the entire bands for a column relative to the DNA Mass Marker are indicated below the corresponding lane. Lanes that did not have any detectable bands over background are reported as not detectable (ND).
    Figure Legend Snippet: ZIKV RT-LAMP sensitivity for ZIKV. Sensitivity assessment of ZIKV RT-LAMP using serial dilutions of ZIKV PCR Standard (Robert Koch Institute) from 4 × 10 5 genome copies/reaction to 1 copies/reaction as visualized by addition of SYBR Green I by eye (upper panel), green fluorescence (middle panel), or gel electrophoresis (bottom panel). Lane M: Low DNA Mass Marker (ThermoFisher Scientific); NTC: No template control (negative control). Relative density (RD) of the entire bands for a column relative to the DNA Mass Marker are indicated below the corresponding lane. Lanes that did not have any detectable bands over background are reported as not detectable (ND).

    Techniques Used: Polymerase Chain Reaction, SYBR Green Assay, Fluorescence, Nucleic Acid Electrophoresis, Marker, Negative Control

    ZIKV RT-LAMP specificity for ZIKV. Specificity assessment of ZIKV RT-LAMP in ZIKV or DENV infected whole cell lysates (WCL) or cell culture supernatants (SN) as visualized by the addition of SYBR Green I by eye (upper panel), green fluorescence (middle panel), or gel electrophoresis (bottom panel). Lane M: Low DNA Mass Marker (ThermoFisher Scientific); NTC: No template control (negative control); + : ZIKV PCR Standard (Robert Koch Institute; positive control).
    Figure Legend Snippet: ZIKV RT-LAMP specificity for ZIKV. Specificity assessment of ZIKV RT-LAMP in ZIKV or DENV infected whole cell lysates (WCL) or cell culture supernatants (SN) as visualized by the addition of SYBR Green I by eye (upper panel), green fluorescence (middle panel), or gel electrophoresis (bottom panel). Lane M: Low DNA Mass Marker (ThermoFisher Scientific); NTC: No template control (negative control); + : ZIKV PCR Standard (Robert Koch Institute; positive control).

    Techniques Used: Infection, Cell Culture, SYBR Green Assay, Fluorescence, Nucleic Acid Electrophoresis, Marker, Negative Control, Polymerase Chain Reaction, Positive Control

    ZIKV and Actin detection in mosquitos. ( A ) qRT-PCR of ZIKV RNA normalized to total RNA for mock (n = 5) or ZIKV (n = 5) infected mosquitos. ( B , C ) Single mosquitos were infected with either mock, ZIKV, or DENV and subjected to a ZIKV ( B ) or Aedes aegypti Actin ( C ) specific RT-LAMP reaction then visualized by addition of SYBR Green I by eye (upper panel), green fluorescence (middle panel), or gel electrophoresis (bottom panel). Lane M: Low DNA Mass Marker (ThermoFisher Scientific); NTC: No template control (negative control). Relative density (RD) of the entire bands for a column relative to the DNA Mass Marker are indicated below the corresponding lane. Lanes that did not have any detectable bands over background are reported as not detectable (ND). ( D ) Detection of ZIKV or ribosomal s17 by qRT-PCR. Data shown as mean ± standard deviation.
    Figure Legend Snippet: ZIKV and Actin detection in mosquitos. ( A ) qRT-PCR of ZIKV RNA normalized to total RNA for mock (n = 5) or ZIKV (n = 5) infected mosquitos. ( B , C ) Single mosquitos were infected with either mock, ZIKV, or DENV and subjected to a ZIKV ( B ) or Aedes aegypti Actin ( C ) specific RT-LAMP reaction then visualized by addition of SYBR Green I by eye (upper panel), green fluorescence (middle panel), or gel electrophoresis (bottom panel). Lane M: Low DNA Mass Marker (ThermoFisher Scientific); NTC: No template control (negative control). Relative density (RD) of the entire bands for a column relative to the DNA Mass Marker are indicated below the corresponding lane. Lanes that did not have any detectable bands over background are reported as not detectable (ND). ( D ) Detection of ZIKV or ribosomal s17 by qRT-PCR. Data shown as mean ± standard deviation.

    Techniques Used: Quantitative RT-PCR, Infection, SYBR Green Assay, Fluorescence, Nucleic Acid Electrophoresis, Marker, Negative Control, Standard Deviation

    RT-LAMP detection of ZIKV. ( A ) ZIKA RT-LAMP amplification of ZIKV PCR Standard (ZIKV; Robert Koch Institute) but not no template control (NTC; negative control) as visualized by addition of SYBR Green I (SYBR) by eye (upper panel), green fluorescence (middle panel), or gel electrophoresis (bottom panel). Lane M: Low DNA Mass Marker (ThermoFisher Scientific). ( B ) All primers (All) are required for effective LAMP reaction. Reactions without FIP and BIP (-FIP/BIP) or FL and BL (-FL/BL) resulted in a negative RT-LAMP reaction.
    Figure Legend Snippet: RT-LAMP detection of ZIKV. ( A ) ZIKA RT-LAMP amplification of ZIKV PCR Standard (ZIKV; Robert Koch Institute) but not no template control (NTC; negative control) as visualized by addition of SYBR Green I (SYBR) by eye (upper panel), green fluorescence (middle panel), or gel electrophoresis (bottom panel). Lane M: Low DNA Mass Marker (ThermoFisher Scientific). ( B ) All primers (All) are required for effective LAMP reaction. Reactions without FIP and BIP (-FIP/BIP) or FL and BL (-FL/BL) resulted in a negative RT-LAMP reaction.

    Techniques Used: Amplification, Polymerase Chain Reaction, Negative Control, SYBR Green Assay, Fluorescence, Nucleic Acid Electrophoresis, Marker

    3) Product Images from "An Adaptable Two-Color Flow Cytometric Assay to Quantitate the Invasion of Erythrocytes by Plasmodium falciparum Parasites"

    Article Title: An Adaptable Two-Color Flow Cytometric Assay to Quantitate the Invasion of Erythrocytes by Plasmodium falciparum Parasites

    Journal: Cytometry

    doi: 10.1002/cyto.a.20972

    Staining of pRBC with fluorescent DNA-binding dyes. pRBC were detected by flow cytometry after staining with either 10 μg/mL EB, 2 μM Hoechst 33342, or 1:5,000 SYBR Green I. a : Uninfected RBC and pRBC were directly stained with the DNA dyes. b : Uninfected RBC and pRBC were fixed, permeabilized, and treated with RNase before staining with the DNA dyes.
    Figure Legend Snippet: Staining of pRBC with fluorescent DNA-binding dyes. pRBC were detected by flow cytometry after staining with either 10 μg/mL EB, 2 μM Hoechst 33342, or 1:5,000 SYBR Green I. a : Uninfected RBC and pRBC were directly stained with the DNA dyes. b : Uninfected RBC and pRBC were fixed, permeabilized, and treated with RNase before staining with the DNA dyes.

    Techniques Used: Staining, Binding Assay, Flow Cytometry, Cytometry, SYBR Green Assay

    Accuracy of flow cytometry in determining parasitemia. Correlation between parasitemia determined by flow cytometry, using either Hoechst 33342 (direct staining) or SYBR Green I (staining post RNase treatment), and by light microscopy, using Giemsa staining, for a serial dilution of a Dd2 parasite culture. Standard error bars are represented on the horizontal axis for microscopy counts and vertical axis for flow cytometry counts.
    Figure Legend Snippet: Accuracy of flow cytometry in determining parasitemia. Correlation between parasitemia determined by flow cytometry, using either Hoechst 33342 (direct staining) or SYBR Green I (staining post RNase treatment), and by light microscopy, using Giemsa staining, for a serial dilution of a Dd2 parasite culture. Standard error bars are represented on the horizontal axis for microscopy counts and vertical axis for flow cytometry counts.

    Techniques Used: Flow Cytometry, Cytometry, Staining, SYBR Green Assay, Light Microscopy, Serial Dilution, Microscopy

    Invasion phenotypes of three laboratory strains of P. falciparum . Unlabeled pRBC were incubated with CFDA-SE- or DDAO-SE-labeled RBC, at an unlabeled-to-labeled ratio of 1:2 and a starting parasitemia of 1%. After 48 h, parasites were stained with Hoechst 33342 or SYBR Green I, respectively, and final parasitemia in the target population was determined by gating on the fluorescently labeled RBC population using flow cytometry. Invasion efficiencies were determined as a percentage of the final parasitemia of a mock-treated-labeled positive control RBC group.
    Figure Legend Snippet: Invasion phenotypes of three laboratory strains of P. falciparum . Unlabeled pRBC were incubated with CFDA-SE- or DDAO-SE-labeled RBC, at an unlabeled-to-labeled ratio of 1:2 and a starting parasitemia of 1%. After 48 h, parasites were stained with Hoechst 33342 or SYBR Green I, respectively, and final parasitemia in the target population was determined by gating on the fluorescently labeled RBC population using flow cytometry. Invasion efficiencies were determined as a percentage of the final parasitemia of a mock-treated-labeled positive control RBC group.

    Techniques Used: Incubation, Labeling, Staining, SYBR Green Assay, Flow Cytometry, Cytometry, Positive Control

    P. falciparum invasion in labeled cells. Target RBC labeled with fluorescent dyes CFDA-SE or DDAO-SE were coincubated with unlabeled P. falciparum Dd2 strain cultures, containing a mix of uninfected and pRBC, for 48 h under standard P. falciparum culture conditions. Cultures were then harvested and stained with Hoechst 33342 or SYBR Green I, respectively. a : Cultures were observed by confocal microscopy (first 4 panels) and flow cytometry (last panel on the right). In the microscopy pictures, yellow (Hoechst 33342) and blue (SYBR Green I) arrows point to parasites detected inside unlabeled RBC, whereas white arrows point to parasites detected inside fluorescently labeled RBC. In the dotplot representation of the data generated by flow cytometry, four populations can be readily distinguished: unlabeled, uninfected RBC (lower left); labeled, uninfected RBC (lower right); unlabeled, infected RBC (upper left); and labeled, infected RBCs (upper right). b : Effect of the starting parasitemia on the final parasitemia in labeled RBC. Parasitemia of DDAO-SE-labeled RBC was determined by SYBR Green I staining using flow cytometry, for different starting parasitemia using donor unlabeled population with increasing parasitemia. c : Effect of the unlabeled to labeled RBC ratio on the final parasitemia in labeled RBC. Parasitemia of DDAO-SE-labeled RBC was determined by SYBR Green I staining using flow cytometry, in a mixed unlabeled donor pRBC/label target RBC culture with a volume of 100 μL and a starting parasitemia of 1%.
    Figure Legend Snippet: P. falciparum invasion in labeled cells. Target RBC labeled with fluorescent dyes CFDA-SE or DDAO-SE were coincubated with unlabeled P. falciparum Dd2 strain cultures, containing a mix of uninfected and pRBC, for 48 h under standard P. falciparum culture conditions. Cultures were then harvested and stained with Hoechst 33342 or SYBR Green I, respectively. a : Cultures were observed by confocal microscopy (first 4 panels) and flow cytometry (last panel on the right). In the microscopy pictures, yellow (Hoechst 33342) and blue (SYBR Green I) arrows point to parasites detected inside unlabeled RBC, whereas white arrows point to parasites detected inside fluorescently labeled RBC. In the dotplot representation of the data generated by flow cytometry, four populations can be readily distinguished: unlabeled, uninfected RBC (lower left); labeled, uninfected RBC (lower right); unlabeled, infected RBC (upper left); and labeled, infected RBCs (upper right). b : Effect of the starting parasitemia on the final parasitemia in labeled RBC. Parasitemia of DDAO-SE-labeled RBC was determined by SYBR Green I staining using flow cytometry, for different starting parasitemia using donor unlabeled population with increasing parasitemia. c : Effect of the unlabeled to labeled RBC ratio on the final parasitemia in labeled RBC. Parasitemia of DDAO-SE-labeled RBC was determined by SYBR Green I staining using flow cytometry, in a mixed unlabeled donor pRBC/label target RBC culture with a volume of 100 μL and a starting parasitemia of 1%.

    Techniques Used: Labeling, Staining, SYBR Green Assay, Confocal Microscopy, Flow Cytometry, Cytometry, Microscopy, Generated, Infection

    4) Product Images from "Counting Viruses and Bacteria in Photosynthetic Microbial Mats"

    Article Title: Counting Viruses and Bacteria in Photosynthetic Microbial Mats

    Journal: Applied and Environmental Microbiology

    doi: 10.1128/AEM.02863-14

    Cytogram of viruses from photosynthetic microbial mat samples using flow cytometry after staining with nucleic acid-specific dye SYBR green I (A) and from control sample without viruses (B). Green fluorescence (V1 and V2) allows the distinction of two virus clusters.
    Figure Legend Snippet: Cytogram of viruses from photosynthetic microbial mat samples using flow cytometry after staining with nucleic acid-specific dye SYBR green I (A) and from control sample without viruses (B). Green fluorescence (V1 and V2) allows the distinction of two virus clusters.

    Techniques Used: Flow Cytometry, Cytometry, Staining, SYBR Green Assay, Fluorescence

    5) Product Images from "One-Pot Reverse Transcriptional Loop-Mediated Isothermal Amplification (RT-LAMP) for Detecting MERS-CoV"

    Article Title: One-Pot Reverse Transcriptional Loop-Mediated Isothermal Amplification (RT-LAMP) for Detecting MERS-CoV

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2016.02166

    (A) The RT-LAMP detection results based on different ratios of Bst polymerase and M-MLV reverse transcriptase concentration (100 unit). Concentration of Bst polymerse is 1, 2, 4, 6, and 8 units. (B) Effect of reaction time for RT-LAMP reaction. DNA ladder-like pattern was confirmed by 2% agarose gel electrophoresis. LAMP products detected by adding 1,000X SYBR-green I when the reaction was completed (UV/on and off). Land M: 100 bp DNA ladders; - depicts tube used as negative control, non-template; + indicates the results with template.
    Figure Legend Snippet: (A) The RT-LAMP detection results based on different ratios of Bst polymerase and M-MLV reverse transcriptase concentration (100 unit). Concentration of Bst polymerse is 1, 2, 4, 6, and 8 units. (B) Effect of reaction time for RT-LAMP reaction. DNA ladder-like pattern was confirmed by 2% agarose gel electrophoresis. LAMP products detected by adding 1,000X SYBR-green I when the reaction was completed (UV/on and off). Land M: 100 bp DNA ladders; - depicts tube used as negative control, non-template; + indicates the results with template.

    Techniques Used: Concentration Assay, Agarose Gel Electrophoresis, SYBR Green Assay, Negative Control

    6) Product Images from "Transcriptome analysis of functional differentiation between haploid and diploid cells of Emiliania huxleyi, a globally significant photosynthetic calcifying cell"

    Article Title: Transcriptome analysis of functional differentiation between haploid and diploid cells of Emiliania huxleyi, a globally significant photosynthetic calcifying cell

    Journal: Genome Biology

    doi: 10.1186/gb-2009-10-10-r114

    Cell cycle changes during the day-night cycle of harvesting. Example DNA content histograms of nuclear extracts taken from 1N cultures at different times are shown. The time point at 15 h on day 1 is not shown but had a similar distribution to that at 19 h on day 1 and 15 h30 on day 2. RNA was not collected at 15 h30 on day 2, but nuclear extracts (shown here), flow cytometric profiles, and Fv/Fm confirmed cells had returned to the same state after a complete diel cycle. Extracted nuclei were stained with Sybr Green I and analyzed by flow cytometry.
    Figure Legend Snippet: Cell cycle changes during the day-night cycle of harvesting. Example DNA content histograms of nuclear extracts taken from 1N cultures at different times are shown. The time point at 15 h on day 1 is not shown but had a similar distribution to that at 19 h on day 1 and 15 h30 on day 2. RNA was not collected at 15 h30 on day 2, but nuclear extracts (shown here), flow cytometric profiles, and Fv/Fm confirmed cells had returned to the same state after a complete diel cycle. Extracted nuclei were stained with Sybr Green I and analyzed by flow cytometry.

    Techniques Used: Flow Cytometry, Staining, SYBR Green Assay, Cytometry

    7) Product Images from "Inducing controlled cell cycle arrest and re-entry during asexual proliferation of Plasmodium falciparum malaria parasites"

    Article Title: Inducing controlled cell cycle arrest and re-entry during asexual proliferation of Plasmodium falciparum malaria parasites

    Journal: Scientific Reports

    doi: 10.1038/s41598-018-34964-w

    DFMO arrest and putrescine reversal associated to G1/S cell cycle control in intraerythrocytic P. falciparum parasites. In all instances, ■ refers to control parasites, ▼ refers to DFMO-treated parasites, and ● refers to putrescine-reversed parasites. ( a ) DFMO dose-response curves of asexual P . falciparum 3D7 proliferation over 96 h (initiated with ring-stage parasites, 1% haematocrit, 1% parasitaemia) at 37 °C in the absence or presence of 2 mM putrescine. Proliferation is expressed relative to untreated controls, with data averaged from n = 6 biological replicates and shown ± S.E. ( b ) Synchronised P . falciparum 3D7 cultures were treated with DFMO alone (IC 90 ) or with putrescine (2 mM, after 24 h DFMO pressure) and parasitaemia monitored over 96 h with SYBR Green I fluorescence (10 000 infected erythrocytes counted). * P
    Figure Legend Snippet: DFMO arrest and putrescine reversal associated to G1/S cell cycle control in intraerythrocytic P. falciparum parasites. In all instances, ■ refers to control parasites, ▼ refers to DFMO-treated parasites, and ● refers to putrescine-reversed parasites. ( a ) DFMO dose-response curves of asexual P . falciparum 3D7 proliferation over 96 h (initiated with ring-stage parasites, 1% haematocrit, 1% parasitaemia) at 37 °C in the absence or presence of 2 mM putrescine. Proliferation is expressed relative to untreated controls, with data averaged from n = 6 biological replicates and shown ± S.E. ( b ) Synchronised P . falciparum 3D7 cultures were treated with DFMO alone (IC 90 ) or with putrescine (2 mM, after 24 h DFMO pressure) and parasitaemia monitored over 96 h with SYBR Green I fluorescence (10 000 infected erythrocytes counted). * P

    Techniques Used: SYBR Green Assay, Fluorescence, Infection

    8) Product Images from "Targeted Sorting of Single Virus-Infected Cells of the Coccolithophore Emiliania huxleyi"

    Article Title: Targeted Sorting of Single Virus-Infected Cells of the Coccolithophore Emiliania huxleyi

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0022520

    Representative flow cytometry plots showing E. huxleyi cells, inoculated and non-inoculated with viruses (20 hours PI) (A) without fluorescence dye, or stained with the fluorescence dyes (B) lipid-specific dye FM 1-43, (C) CM-H 2 DCFDA for detection of accumulated Reactive Oxygen Species in cells and (D) DNA dye SYBR Green I. Infected and non-infected cells were discriminated on the basis of their red autofluorescence (610 nm) or the green fluorescence (522 nm) of SYBR Green I and CM-H 2 DCFDA versus side scatter, green dye fluorescence or orange lipid dye FM 1-43 fluorescence (488 nm).
    Figure Legend Snippet: Representative flow cytometry plots showing E. huxleyi cells, inoculated and non-inoculated with viruses (20 hours PI) (A) without fluorescence dye, or stained with the fluorescence dyes (B) lipid-specific dye FM 1-43, (C) CM-H 2 DCFDA for detection of accumulated Reactive Oxygen Species in cells and (D) DNA dye SYBR Green I. Infected and non-infected cells were discriminated on the basis of their red autofluorescence (610 nm) or the green fluorescence (522 nm) of SYBR Green I and CM-H 2 DCFDA versus side scatter, green dye fluorescence or orange lipid dye FM 1-43 fluorescence (488 nm).

    Techniques Used: Flow Cytometry, Cytometry, Fluorescence, Staining, SYBR Green Assay, Infection

    9) Product Images from "Monitoring of Dynamic Microbiological Processes Using Real-Time Flow Cytometry"

    Article Title: Monitoring of Dynamic Microbiological Processes Using Real-Time Flow Cytometry

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0080117

    Temperature induced cell damage in a river water sample heated from 35 - 70 °C. ( A ) Density plot of the raw water stained with SYBR Green I and propidium iodide, showing the regions where “intact” and “damaged” cells are located, as well as the clusters of low (LNA; blue) and high (HNA; green) nucleic acid content bacteria. ( B ) Raw RT-FCM data of green fluorescence (ex. 488 nm/em. 530 nm) intensity over time for the intact cells (from panel A). ( C ) Processed data (1-minute intervals) showing the decrease in intact cell concentrations during the experiment directly linked to the increase in temperature. ( D ) Relative changes of the concentrations of the LNA (blue triangles) and HNA (green diamonds) bacterial fractions against temperature, showing a higher sensitivity to temperature in the HNA population.
    Figure Legend Snippet: Temperature induced cell damage in a river water sample heated from 35 - 70 °C. ( A ) Density plot of the raw water stained with SYBR Green I and propidium iodide, showing the regions where “intact” and “damaged” cells are located, as well as the clusters of low (LNA; blue) and high (HNA; green) nucleic acid content bacteria. ( B ) Raw RT-FCM data of green fluorescence (ex. 488 nm/em. 530 nm) intensity over time for the intact cells (from panel A). ( C ) Processed data (1-minute intervals) showing the decrease in intact cell concentrations during the experiment directly linked to the increase in temperature. ( D ) Relative changes of the concentrations of the LNA (blue triangles) and HNA (green diamonds) bacterial fractions against temperature, showing a higher sensitivity to temperature in the HNA population.

    Techniques Used: Staining, SYBR Green Assay, Fluorescence

    10) Product Images from "Application of a loop-mediated isothermal amplification (LAMP) assay targeting cox1 gene for the detection of Clonorchis sinensis in human fecal samples"

    Article Title: Application of a loop-mediated isothermal amplification (LAMP) assay targeting cox1 gene for the detection of Clonorchis sinensis in human fecal samples

    Journal: PLoS Neglected Tropical Diseases

    doi: 10.1371/journal.pntd.0005995

    Sensitivity of the loop-mediated isothermal amplification (LAMP) assay for the detection of Clonorchis sinensis eggs in feces experimentally spiked with a known number of eggs in ten-fold serial dilutions from 10,000 eggs (lane 1) to 1 egg (lane 5). (A) Naked eye detection of LAMP products using SYBR Green I. A green color indicates a positive reaction, and an orange color indicates a negative reaction. (B) Fluorescence of LAMP products after using SYBR Green I followed by detection under UV light. (C) Agarose gel electrophoresis of LAMP products followed by ethidium bromide staining and detection under UV light. Values in the left are in base pairs. Lane 6, negative stool DNA; lane M, molecular marker.
    Figure Legend Snippet: Sensitivity of the loop-mediated isothermal amplification (LAMP) assay for the detection of Clonorchis sinensis eggs in feces experimentally spiked with a known number of eggs in ten-fold serial dilutions from 10,000 eggs (lane 1) to 1 egg (lane 5). (A) Naked eye detection of LAMP products using SYBR Green I. A green color indicates a positive reaction, and an orange color indicates a negative reaction. (B) Fluorescence of LAMP products after using SYBR Green I followed by detection under UV light. (C) Agarose gel electrophoresis of LAMP products followed by ethidium bromide staining and detection under UV light. Values in the left are in base pairs. Lane 6, negative stool DNA; lane M, molecular marker.

    Techniques Used: Amplification, Lamp Assay, SYBR Green Assay, Fluorescence, Agarose Gel Electrophoresis, Staining, Marker

    Specificity of the loop-mediated isothermal amplification (LAMP) assay for the detection of Clonorchis sinensis genomic DNA. (A) Naked eye detection of LAMP products using SYBR Green I. A green color indicates a positive reaction, and an orange color indicates a negative reaction. (B) Fluorescence of LAMP products after using SYBR Green I followed by detection under UV light. (C) Agarose gel electrophoresis of LAMP products followed by ethidium bromide staining and detection under UV light. Values in the left are in base pairs. Lane 1, Clonorchis sinensis ; lane 2, Metagonimus yokogawai ; lane 3, Opisthorchis viverrini ; lane 4, Fasciola gigantica ; lane 5, Spirometra erinacei ; lane 6, Diphyllobothrium latum ; lane 7, Ascaris lumbricoides ; lane 8, Ascaris suum ; lane 9, Necator americanus ; lane 10, Trichuris trichiura ; lane 11, Cryptosporidium parvum ; lane 12, Entamoeba histolytica ; lane 13, Giardia lamblia ; lane 14, Escherichia coli ; lane 15, non-template control; M, molecular marker.
    Figure Legend Snippet: Specificity of the loop-mediated isothermal amplification (LAMP) assay for the detection of Clonorchis sinensis genomic DNA. (A) Naked eye detection of LAMP products using SYBR Green I. A green color indicates a positive reaction, and an orange color indicates a negative reaction. (B) Fluorescence of LAMP products after using SYBR Green I followed by detection under UV light. (C) Agarose gel electrophoresis of LAMP products followed by ethidium bromide staining and detection under UV light. Values in the left are in base pairs. Lane 1, Clonorchis sinensis ; lane 2, Metagonimus yokogawai ; lane 3, Opisthorchis viverrini ; lane 4, Fasciola gigantica ; lane 5, Spirometra erinacei ; lane 6, Diphyllobothrium latum ; lane 7, Ascaris lumbricoides ; lane 8, Ascaris suum ; lane 9, Necator americanus ; lane 10, Trichuris trichiura ; lane 11, Cryptosporidium parvum ; lane 12, Entamoeba histolytica ; lane 13, Giardia lamblia ; lane 14, Escherichia coli ; lane 15, non-template control; M, molecular marker.

    Techniques Used: Amplification, Lamp Assay, SYBR Green Assay, Fluorescence, Agarose Gel Electrophoresis, Staining, Marker

    Sensitivity of the loop-mediated isothermal amplification (LAMP) assay and PCR for the detection of Clonorchis sinensis genomic DNA. Ten-fold serial dilutions starting from 1 ng of genomic DNA (lane 1) down to 1 fg (lane 7) were tested. (A) Naked eye detection of LAMP products using SYBR Green I. A green color indicates a positive reaction, and an orange color indicates a negative reaction. (B) Fluorescence of LAMP products after using SYBR Green I followed by detection under UV light. (C) Agarose gel electrophoresis of LAMP products followed by ethidium bromide staining and detection under UV light. (D) PCR with outer primers F3 and B3. Values in the left are in base pairs. Lane 8, non-template control (NTC); lane M, molecular marker.
    Figure Legend Snippet: Sensitivity of the loop-mediated isothermal amplification (LAMP) assay and PCR for the detection of Clonorchis sinensis genomic DNA. Ten-fold serial dilutions starting from 1 ng of genomic DNA (lane 1) down to 1 fg (lane 7) were tested. (A) Naked eye detection of LAMP products using SYBR Green I. A green color indicates a positive reaction, and an orange color indicates a negative reaction. (B) Fluorescence of LAMP products after using SYBR Green I followed by detection under UV light. (C) Agarose gel electrophoresis of LAMP products followed by ethidium bromide staining and detection under UV light. (D) PCR with outer primers F3 and B3. Values in the left are in base pairs. Lane 8, non-template control (NTC); lane M, molecular marker.

    Techniques Used: Amplification, Lamp Assay, Polymerase Chain Reaction, SYBR Green Assay, Fluorescence, Agarose Gel Electrophoresis, Staining, Marker

    11) Product Images from "A high-throughput qPCR system for simultaneous quantitative detection of dairy Lactococcus lactis and Leuconostoc bacteriophages"

    Article Title: A high-throughput qPCR system for simultaneous quantitative detection of dairy Lactococcus lactis and Leuconostoc bacteriophages

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0174223

    Performance of SYBR Green I and EvaGreen detection chemistries during qPCR assays. (A) Standard curves generated from amplification of serially diluted L . lactis phage P220 genome detected with the corresponding chemistries; (B) the corresponding performance parameters; and (C) amplification plots detected with SYBR Green I (brown) and EvaGreen (green).
    Figure Legend Snippet: Performance of SYBR Green I and EvaGreen detection chemistries during qPCR assays. (A) Standard curves generated from amplification of serially diluted L . lactis phage P220 genome detected with the corresponding chemistries; (B) the corresponding performance parameters; and (C) amplification plots detected with SYBR Green I (brown) and EvaGreen (green).

    Techniques Used: SYBR Green Assay, Real-time Polymerase Chain Reaction, Generated, Amplification

    12) Product Images from "Frequent fusion and fission of plant mitochondria with unequal nucleoid distribution"

    Article Title: Frequent fusion and fission of plant mitochondria with unequal nucleoid distribution

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    doi: 10.1073/pnas.0401077101

    Mitochondria and mitochondrial nucleoids in tobacco suspension cultured cells (BY-2). ( a ) Mitochondria stained by MitoTracker ( Left ) and mitochondrial nucleoids stained by Sybr Green I ( Right ). In the merged images ( Center ), the nucleoids appear yellow because of the combination of red and green fluorescence. Arrowheads indicate mitochondria that have no visible nucleoids. ( b ) Views of three mitochondria at 0.5-μm intervals along the optical axis. Yellow nucleoids are visible in the lower two mitochondria but not in the top mitochondria. ( c and d ) Time series of fission of mitochondria with nucleoids shown at 10-sec intervals. (Scale bars, 2 μm.) Movies 2–4.
    Figure Legend Snippet: Mitochondria and mitochondrial nucleoids in tobacco suspension cultured cells (BY-2). ( a ) Mitochondria stained by MitoTracker ( Left ) and mitochondrial nucleoids stained by Sybr Green I ( Right ). In the merged images ( Center ), the nucleoids appear yellow because of the combination of red and green fluorescence. Arrowheads indicate mitochondria that have no visible nucleoids. ( b ) Views of three mitochondria at 0.5-μm intervals along the optical axis. Yellow nucleoids are visible in the lower two mitochondria but not in the top mitochondria. ( c and d ) Time series of fission of mitochondria with nucleoids shown at 10-sec intervals. (Scale bars, 2 μm.) Movies 2–4.

    Techniques Used: Cell Culture, Staining, SYBR Green Assay, Fluorescence, Size-exclusion Chromatography

    13) Product Images from "Effectiveness of Stevia Rebaudiana Whole Leaf Extract Against the Various Morphological Forms of Borrelia Burgdorferi in Vitro"

    Article Title: Effectiveness of Stevia Rebaudiana Whole Leaf Extract Against the Various Morphological Forms of Borrelia Burgdorferi in Vitro

    Journal: European Journal of Microbiology & Immunology

    doi: 10.1556/1886.2015.00031

    Susceptibility of log phase (5 days) and stationary phase (8 days) Borrelia burgdorferi to antimicrobial agents after a three-day treatment determined by (A) SYBR Green I/PI assay, (B) direct counting of live and dead cells stained using a mixture of SYBR Green I and propidium iodide using fluorescent microscopy. (C) Representative live/dead images of log and stationary phase B. burgdorferi to antimicrobial agents taken at 200× magnification (Scale bar – 100 μm). Doxycycline (DoxC) was used as a positive control. 1× PBS and 25% alcohol were used as negative controls, respectively. All antibiotics individually as well as in combination were used at a concentration of 10 μg/ml. Stevia A was used at a concentration of 1.2 μg/ml. n = 3 ± SD, * p ≤ 0.05, ** p ≤ 0.01 compared to the control. n = 3 ± SD, + p ≤ 0.05, ++ p ≤ 0.01 (doxycycline compared to Stevia A). n = 3 ± SD, * p ≤ 0.05, ** p ≤ 0.01 (doxycycline compared to Stevia A). Abbreviations: doxycycline – DoxC, cefoperazone – CefP, daptomycin – DapM
    Figure Legend Snippet: Susceptibility of log phase (5 days) and stationary phase (8 days) Borrelia burgdorferi to antimicrobial agents after a three-day treatment determined by (A) SYBR Green I/PI assay, (B) direct counting of live and dead cells stained using a mixture of SYBR Green I and propidium iodide using fluorescent microscopy. (C) Representative live/dead images of log and stationary phase B. burgdorferi to antimicrobial agents taken at 200× magnification (Scale bar – 100 μm). Doxycycline (DoxC) was used as a positive control. 1× PBS and 25% alcohol were used as negative controls, respectively. All antibiotics individually as well as in combination were used at a concentration of 10 μg/ml. Stevia A was used at a concentration of 1.2 μg/ml. n = 3 ± SD, * p ≤ 0.05, ** p ≤ 0.01 compared to the control. n = 3 ± SD, + p ≤ 0.05, ++ p ≤ 0.01 (doxycycline compared to Stevia A). n = 3 ± SD, * p ≤ 0.05, ** p ≤ 0.01 (doxycycline compared to Stevia A). Abbreviations: doxycycline – DoxC, cefoperazone – CefP, daptomycin – DapM

    Techniques Used: SYBR Green Assay, Staining, Microscopy, Positive Control, Concentration Assay

    Representative live/dead images of Borrelia biofilms treated with different antimicrobial agents followed by staining with SYBR Green I and PI dye mixture taken at 200× magnification. Doxycycline (DoxC) was used as a positive control. 1× sterile PBS and 25% alcohol were used as negative controls, respectively. All antibiotics individually as well as in combination were used at a concentration of 10 μg/ml. Stevia A was used at a concentration of 1.2 μg/ml. Scale bar –100 μm. Abbreviations: doxycycline – DoxC, cefoperazone – CefP, daptomycin – DapM
    Figure Legend Snippet: Representative live/dead images of Borrelia biofilms treated with different antimicrobial agents followed by staining with SYBR Green I and PI dye mixture taken at 200× magnification. Doxycycline (DoxC) was used as a positive control. 1× sterile PBS and 25% alcohol were used as negative controls, respectively. All antibiotics individually as well as in combination were used at a concentration of 10 μg/ml. Stevia A was used at a concentration of 1.2 μg/ml. Scale bar –100 μm. Abbreviations: doxycycline – DoxC, cefoperazone – CefP, daptomycin – DapM

    Techniques Used: Staining, SYBR Green Assay, Positive Control, Concentration Assay

    A 7-day and 14-day subculture of 8-day-old Borrelia burgdorferi stationary phase culture treated with antimicrobial agents determined by (A) SYBR Green I/PI assay and direct counting of live and dead cells stained using a mixture of SYBR Green I and propidium iodide using fluorescent microscopy. (B) Representative live/dead images of a 7-day-old subculture on stationary phase B. burgdorferi treated with antimicrobial agents taken at 200× magnification (Scale bar – 100 μm). Doxycycline (DoxC) was used as a positive control. 1× PBS and 25% alcohol were used as negative controls respectively. n = 3 ± SD, * p ≤ 0.05, ** p ≤ 0.01 compared to the control. n = 3 ± SD, + p ≤ 0.05, ++ p
    Figure Legend Snippet: A 7-day and 14-day subculture of 8-day-old Borrelia burgdorferi stationary phase culture treated with antimicrobial agents determined by (A) SYBR Green I/PI assay and direct counting of live and dead cells stained using a mixture of SYBR Green I and propidium iodide using fluorescent microscopy. (B) Representative live/dead images of a 7-day-old subculture on stationary phase B. burgdorferi treated with antimicrobial agents taken at 200× magnification (Scale bar – 100 μm). Doxycycline (DoxC) was used as a positive control. 1× PBS and 25% alcohol were used as negative controls respectively. n = 3 ± SD, * p ≤ 0.05, ** p ≤ 0.01 compared to the control. n = 3 ± SD, + p ≤ 0.05, ++ p

    Techniques Used: SYBR Green Assay, Staining, Microscopy, Positive Control

    14) Product Images from "Evaluation of a Direct Reverse Transcription Loop-Mediated Isothermal Amplification Method without RNA Extraction for the Detection of Human Enterovirus 71 Subgenotype C4 in Nasopharyngeal Swab Specimens"

    Article Title: Evaluation of a Direct Reverse Transcription Loop-Mediated Isothermal Amplification Method without RNA Extraction for the Detection of Human Enterovirus 71 Subgenotype C4 in Nasopharyngeal Swab Specimens

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0052486

    Visual detection of amplified LAMP products using SYBR green I. Addition of 1 µl of diluted SYBR green I to the reaction tube after LAMP reaction enables visible analysis of the results under natural light ( Figure 1A ) or UV irradiation ( Figure 1B ). The color changes from orange (negative reaction) to green (positive reaction) ( Figure 1A ) and bright fluorescence indicates a positive reaction ( Figure 1B ).
    Figure Legend Snippet: Visual detection of amplified LAMP products using SYBR green I. Addition of 1 µl of diluted SYBR green I to the reaction tube after LAMP reaction enables visible analysis of the results under natural light ( Figure 1A ) or UV irradiation ( Figure 1B ). The color changes from orange (negative reaction) to green (positive reaction) ( Figure 1A ) and bright fluorescence indicates a positive reaction ( Figure 1B ).

    Techniques Used: Amplification, SYBR Green Assay, Irradiation, Fluorescence

    15) Product Images from "Exclusion of small terminase mediated DNA threading models for genome packaging in bacteriophage T4"

    Article Title: Exclusion of small terminase mediated DNA threading models for genome packaging in bacteriophage T4

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkw184

    The oligomerization domain alone is insufficient for DNA binding. ( A ) Schematic of the oligomerization domain deletion construct. Numbers represent the amino acid positions in the gp16 coding sequence. ( B ) Elution profile of the gp16 oligomerization domain deletion mutant. The void volume is indicated by an arrow. Oligomeric state of the domain was calculated from the elution volume. ( C ) The peak fraction from (B) was electrophoresed on a 4–20% gradient native polyacrylamide gel and the gel was stained with Coomassie blue for protein (lane 1) and SYBR Green I for DNA (lane 2). ( D ) In vitro DNA binding of the oligomerization domain deletion mutant. The purified protein (25–75 μM) was incubated with the 500-bp g16 DNA (5 nM) in a 12 μl reaction mixture for 15 min. Samples were then electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with SYBR Green I. The control lane 1 contained no gp16.
    Figure Legend Snippet: The oligomerization domain alone is insufficient for DNA binding. ( A ) Schematic of the oligomerization domain deletion construct. Numbers represent the amino acid positions in the gp16 coding sequence. ( B ) Elution profile of the gp16 oligomerization domain deletion mutant. The void volume is indicated by an arrow. Oligomeric state of the domain was calculated from the elution volume. ( C ) The peak fraction from (B) was electrophoresed on a 4–20% gradient native polyacrylamide gel and the gel was stained with Coomassie blue for protein (lane 1) and SYBR Green I for DNA (lane 2). ( D ) In vitro DNA binding of the oligomerization domain deletion mutant. The purified protein (25–75 μM) was incubated with the 500-bp g16 DNA (5 nM) in a 12 μl reaction mixture for 15 min. Samples were then electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with SYBR Green I. The control lane 1 contained no gp16.

    Techniques Used: Binding Assay, Construct, Sequencing, Mutagenesis, Staining, SYBR Green Assay, In Vitro, Purification, Incubation

    gp16 binds DNA in vitro . The purified phage T4 gp16 (25–75 μM) was incubated with the substrate DNA (500-bp amplified g16 DNA) (5 nM) in a 12 μl reaction mixture for 15 min. The reaction mixture was electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with Coomassie blue for protein ( A ) and SYBR Green I for DNA ( B ). Control lanes were missing either the gp16 protein (lane 1) or the DNA (lane 5). Position of the shifted DNA band is indicated by a dashed line.
    Figure Legend Snippet: gp16 binds DNA in vitro . The purified phage T4 gp16 (25–75 μM) was incubated with the substrate DNA (500-bp amplified g16 DNA) (5 nM) in a 12 μl reaction mixture for 15 min. The reaction mixture was electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with Coomassie blue for protein ( A ) and SYBR Green I for DNA ( B ). Control lanes were missing either the gp16 protein (lane 1) or the DNA (lane 5). Position of the shifted DNA band is indicated by a dashed line.

    Techniques Used: In Vitro, Purification, Incubation, Amplification, Staining, SYBR Green Assay

    Analysis of in vivo -bound gp16-DNA complex. (A and B) Purified gp16 from phage T4 and related phages (8 μg each) were electrophoresed on a native 4–20% gradient polyacrylamide gel. The same gel was stained with Coomassie blue for protein ( A ) and SYBR Green I for DNA ( B ). The arrow corresponds to a fragment of DNA that was dissociated from RB49 gp16 during electrophoresis (panel B, lane 3). (C and D) Most of the gp16-bound DNA is resistant to Benzonase. The purified T4 and RB49 gp16 proteins (1 mg) were treated with Benzonase (Novagen) overnight at room temperature to digest the loosely-bound DNA fragments. Benzonase was removed by passing the samples through a HiLoad 16/600 Superdex 200 size-exclusion column. The tightly bound nuclease-protected DNA was then released by digesting gp16 with Proteinase K (Thermo Scientific) at 65°C for 30 min. Samples were analyzed on a 4–20% gradient polyacrylamide gel. The gel was stained with Coomassie blue for protein ( C ) and SYBR Green I for DNA ( D ). The positions of the DNA bands released from Proteinase K digestion are marked with arrows (panel D, lanes 4 and 7).
    Figure Legend Snippet: Analysis of in vivo -bound gp16-DNA complex. (A and B) Purified gp16 from phage T4 and related phages (8 μg each) were electrophoresed on a native 4–20% gradient polyacrylamide gel. The same gel was stained with Coomassie blue for protein ( A ) and SYBR Green I for DNA ( B ). The arrow corresponds to a fragment of DNA that was dissociated from RB49 gp16 during electrophoresis (panel B, lane 3). (C and D) Most of the gp16-bound DNA is resistant to Benzonase. The purified T4 and RB49 gp16 proteins (1 mg) were treated with Benzonase (Novagen) overnight at room temperature to digest the loosely-bound DNA fragments. Benzonase was removed by passing the samples through a HiLoad 16/600 Superdex 200 size-exclusion column. The tightly bound nuclease-protected DNA was then released by digesting gp16 with Proteinase K (Thermo Scientific) at 65°C for 30 min. Samples were analyzed on a 4–20% gradient polyacrylamide gel. The gel was stained with Coomassie blue for protein ( C ) and SYBR Green I for DNA ( D ). The positions of the DNA bands released from Proteinase K digestion are marked with arrows (panel D, lanes 4 and 7).

    Techniques Used: In Vivo, Purification, Staining, SYBR Green Assay, Electrophoresis

    Deletion of channel helix α2 did not disrupt DNA binding. ( A ) Schematic of the central domain helix α2 deletion mutants. Numbers represent the amino acid positions in the gp16 coding sequence. Deleted sequences are shown by polylines and the amino acid residues flanking the deletions are shown in blue. ( B ) Elution profiles of mutants Del-1 (magenta), Del-2 (blue) and Del-3 (green). The void volume is indicated by an arrow. The oligomeric state of the gp16 species in peak b of the mutants was calculated from the elution volume. (C and D) The peak gp16 fractions from B were electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with Coomassie blue for protein ( C ) and SYBR Green I for DNA ( D ). ‘a’ and ‘b’ correspond to the fractions from the peaks a and b of the respective mutant. ( E ) In vitro DNA binding of the deletion mutants. The purified proteins (20–60 μM) were incubated with the 500-bp g16 DNA (5 nM) in 12 μl reaction mixture for 15 min. Samples were then electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with SYBR Green I. The control lane 1 contained no gp16 and shifted bands are shown by arrows in lanes 12–16.
    Figure Legend Snippet: Deletion of channel helix α2 did not disrupt DNA binding. ( A ) Schematic of the central domain helix α2 deletion mutants. Numbers represent the amino acid positions in the gp16 coding sequence. Deleted sequences are shown by polylines and the amino acid residues flanking the deletions are shown in blue. ( B ) Elution profiles of mutants Del-1 (magenta), Del-2 (blue) and Del-3 (green). The void volume is indicated by an arrow. The oligomeric state of the gp16 species in peak b of the mutants was calculated from the elution volume. (C and D) The peak gp16 fractions from B were electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with Coomassie blue for protein ( C ) and SYBR Green I for DNA ( D ). ‘a’ and ‘b’ correspond to the fractions from the peaks a and b of the respective mutant. ( E ) In vitro DNA binding of the deletion mutants. The purified proteins (20–60 μM) were incubated with the 500-bp g16 DNA (5 nM) in 12 μl reaction mixture for 15 min. Samples were then electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with SYBR Green I. The control lane 1 contained no gp16 and shifted bands are shown by arrows in lanes 12–16.

    Techniques Used: Binding Assay, Sequencing, Staining, SYBR Green Assay, Mutagenesis, In Vitro, Purification, Incubation

    The positively charged residues of gp16 channel are not essential for DNA binding. ( A ) Schematic of the mutations introduced into the helix α2 of T4 gp16. Numbers represent the amino acid positions in the gp16 coding sequence. Positions of the mutated residues are indicated as black bars in helix α2. ( B ) Elution profiles of WT gp16 (black) and mutants 3M (blue) and 4M (red). The void volume is indicated by an arrow. The hexamer size of gp16 species in peak b of mutant 4M was calculated from the elution volume using the column calibrated with molecular weight standards. (C and D) The peak gp16 fractions from B were electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with Coomassie blue for protein ( C ) and SYBR Green I for DNA ( D ). Lanes 4M-a and 4M-b represent the fractions from peak a and peak b of the mutant 4M. ( E ) In vitro DNA binding of gp16 mutants 3M and 4M. Proteins (20–60 μM) were incubated with the 500-bp g16 DNA (5 nM) in 12 μl reaction mixture for 15 min. Samples were then electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with SYBR Green I. The control lane 1 contained no gp16.
    Figure Legend Snippet: The positively charged residues of gp16 channel are not essential for DNA binding. ( A ) Schematic of the mutations introduced into the helix α2 of T4 gp16. Numbers represent the amino acid positions in the gp16 coding sequence. Positions of the mutated residues are indicated as black bars in helix α2. ( B ) Elution profiles of WT gp16 (black) and mutants 3M (blue) and 4M (red). The void volume is indicated by an arrow. The hexamer size of gp16 species in peak b of mutant 4M was calculated from the elution volume using the column calibrated with molecular weight standards. (C and D) The peak gp16 fractions from B were electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with Coomassie blue for protein ( C ) and SYBR Green I for DNA ( D ). Lanes 4M-a and 4M-b represent the fractions from peak a and peak b of the mutant 4M. ( E ) In vitro DNA binding of gp16 mutants 3M and 4M. Proteins (20–60 μM) were incubated with the 500-bp g16 DNA (5 nM) in 12 μl reaction mixture for 15 min. Samples were then electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with SYBR Green I. The control lane 1 contained no gp16.

    Techniques Used: Binding Assay, Sequencing, Mutagenesis, Molecular Weight, Staining, SYBR Green Assay, In Vitro, Incubation

    16) Product Images from "Calcineurin activation influences muscle phenotype in a muscle-specific fashion"

    Article Title: Calcineurin activation influences muscle phenotype in a muscle-specific fashion

    Journal: BMC Cell Biology

    doi: 10.1186/1471-2121-5-28

    A) SYBR ® Green I stained agarose gel of PCR products following RT-PCR for the MCK-CN* mRNA product. No specific product (~900 bp, arrow) is observed in the DNase-treated RNA samples (first 4 lanes) of either wild-type (WT) or MCK-CN* transgenic (CN*) mouse gastrocnemius. Therefore, there is no contaminating genomic DNA in the RNA samples used for RT-PCR. As expected, RT-PCR of the cDNA (second four lanes) reveals that expression of the transgene is only observed in the MCK-CN* mice. A positive (+) control of tail DNA from an MCK-CN* mouse is shown on the right. B) Western blot of CN* and WT mouse soleus (SOL), diaphragm (DIA), plantaris (PLANT) and gastrocnemius (GAST) muscles for calcineurin. The polyclonal MAb recognized both the endogenous (CN, ~60 kDa) and constitutively active (CN*, ~45 kDa) forms. As expected, muscle from WT muscles did not contain the CN* protein. C) Relative semi-quantitative analysis of CN* mRNA (via RT-PCR) and protein (western blotting) in muscles of MCK-CN* mice. All values were normalized to the same soleus muscle (n = 3 – 6 per group). The asterisk (*) denotes significantly different (p ≤ 0.05) from other muscles.
    Figure Legend Snippet: A) SYBR ® Green I stained agarose gel of PCR products following RT-PCR for the MCK-CN* mRNA product. No specific product (~900 bp, arrow) is observed in the DNase-treated RNA samples (first 4 lanes) of either wild-type (WT) or MCK-CN* transgenic (CN*) mouse gastrocnemius. Therefore, there is no contaminating genomic DNA in the RNA samples used for RT-PCR. As expected, RT-PCR of the cDNA (second four lanes) reveals that expression of the transgene is only observed in the MCK-CN* mice. A positive (+) control of tail DNA from an MCK-CN* mouse is shown on the right. B) Western blot of CN* and WT mouse soleus (SOL), diaphragm (DIA), plantaris (PLANT) and gastrocnemius (GAST) muscles for calcineurin. The polyclonal MAb recognized both the endogenous (CN, ~60 kDa) and constitutively active (CN*, ~45 kDa) forms. As expected, muscle from WT muscles did not contain the CN* protein. C) Relative semi-quantitative analysis of CN* mRNA (via RT-PCR) and protein (western blotting) in muscles of MCK-CN* mice. All values were normalized to the same soleus muscle (n = 3 – 6 per group). The asterisk (*) denotes significantly different (p ≤ 0.05) from other muscles.

    Techniques Used: SYBR Green Assay, Staining, Agarose Gel Electrophoresis, Polymerase Chain Reaction, Reverse Transcription Polymerase Chain Reaction, Transgenic Assay, Expressing, Mouse Assay, Positive Control, Western Blot

    17) Product Images from "Optimization of loop-mediated isothermal amplification (LAMP) assays for the detection of Leishmania DNA in human blood samples"

    Article Title: Optimization of loop-mediated isothermal amplification (LAMP) assays for the detection of Leishmania DNA in human blood samples

    Journal: Acta Tropica

    doi: 10.1016/j.actatropica.2016.06.009

    Agarose gel electrophoresis analysis and SYBR Green I end point detection of LITS-LAMP1 (A,B), and LITS-LAMP2 (C,D). The analysis shows LAMP DNA amplification products of different concentrations amplified from different amounts of L. donovani template DNA, (1) 0.1 ng, (2) 0.01 ng, (3) 1 pg, (4) 0.1 pg. And the amplification of 1 ng template DNA of each of the following Leishmania species: (5) L. major , (6) L. aethiopica , (7) L. tropica , (8) L. infantum chagasi . (9) and (10) No DNA [control], (11–16) Select blood samples obtained by finger prick from volunteers in North Ethiopia. (M) DNA size marker.
    Figure Legend Snippet: Agarose gel electrophoresis analysis and SYBR Green I end point detection of LITS-LAMP1 (A,B), and LITS-LAMP2 (C,D). The analysis shows LAMP DNA amplification products of different concentrations amplified from different amounts of L. donovani template DNA, (1) 0.1 ng, (2) 0.01 ng, (3) 1 pg, (4) 0.1 pg. And the amplification of 1 ng template DNA of each of the following Leishmania species: (5) L. major , (6) L. aethiopica , (7) L. tropica , (8) L. infantum chagasi . (9) and (10) No DNA [control], (11–16) Select blood samples obtained by finger prick from volunteers in North Ethiopia. (M) DNA size marker.

    Techniques Used: Agarose Gel Electrophoresis, SYBR Green Assay, Amplification, Marker

    18) Product Images from "Loop-Mediated Isothermal Amplification Assay for Rapid Detection of Hepatitis C virus"

    Article Title: Loop-Mediated Isothermal Amplification Assay for Rapid Detection of Hepatitis C virus

    Journal: Indian journal of virology : an official organ of Indian Virological Society

    doi: 10.1007/s13337-012-0067-2

    ( a ) Electrophoretic analysis of LAMP amplified products on a 2 % agarose gel; lane M 100-bp DNA ladder marker (Fermentas, Genruler, Germany), lane 1 – 5 LAMP product of Hepatitis C virus , lane 6 LAMP product of positive control, lane 7 LAMP without target RNA, negative control. ( b ) SYBR Green I fluorescent dye-mediated monitoring of HCV LAMP amplification. Visual observation of green fluorescence of DNA binding SYBR Green I under UV light, changed to green in the case of positive amplification, whereas in negative control having no amplification, the original orange color is retained; tubes 1 – 5 LAMP product of HCV, tube 6 LAMP products of positive control, tube 7 LAMP without target RNA. (Color figure online)
    Figure Legend Snippet: ( a ) Electrophoretic analysis of LAMP amplified products on a 2 % agarose gel; lane M 100-bp DNA ladder marker (Fermentas, Genruler, Germany), lane 1 – 5 LAMP product of Hepatitis C virus , lane 6 LAMP product of positive control, lane 7 LAMP without target RNA, negative control. ( b ) SYBR Green I fluorescent dye-mediated monitoring of HCV LAMP amplification. Visual observation of green fluorescence of DNA binding SYBR Green I under UV light, changed to green in the case of positive amplification, whereas in negative control having no amplification, the original orange color is retained; tubes 1 – 5 LAMP product of HCV, tube 6 LAMP products of positive control, tube 7 LAMP without target RNA. (Color figure online)

    Techniques Used: Amplification, Agarose Gel Electrophoresis, Marker, Positive Control, Negative Control, SYBR Green Assay, Fluorescence, Binding Assay

    19) Product Images from "Use of DNA melting simulation software for in silico diagnostic assay design: targeting regions with complex melting curves and confirmation by real-time PCR using intercalating dyes"

    Article Title: Use of DNA melting simulation software for in silico diagnostic assay design: targeting regions with complex melting curves and confirmation by real-time PCR using intercalating dyes

    Journal: BMC Bioinformatics

    doi: 10.1186/1471-2105-8-107

    Comparison of melting profiles obtained using different intercalating dyes and purified Naegleria amplicons . First derivative plots for purified amplified DNA from Naegleria fowleri (red line), Naegleria lovaniensis (blue line) and Naegleria australiensis (black line) melted in the presence of either SYTO9 (A), SYBR Green (B), LC Green I (C) or SYTO59 (D).
    Figure Legend Snippet: Comparison of melting profiles obtained using different intercalating dyes and purified Naegleria amplicons . First derivative plots for purified amplified DNA from Naegleria fowleri (red line), Naegleria lovaniensis (blue line) and Naegleria australiensis (black line) melted in the presence of either SYTO9 (A), SYBR Green (B), LC Green I (C) or SYTO59 (D).

    Techniques Used: Purification, Amplification, SYBR Green Assay

    20) Product Images from "Flow Cytometric Enumeration of Parasitemia in Cultures of Plasmodium falciparum Stained with SYBR Green I and CD235A"

    Article Title: Flow Cytometric Enumeration of Parasitemia in Cultures of Plasmodium falciparum Stained with SYBR Green I and CD235A

    Journal: The Scientific World Journal

    doi: 10.1155/2014/536723

    P. falciparum infected RBCs were stained with SYBR Green I and were then examined using a microscope with (a) DAPI and (b) fluorescence filter. Photographs indicated fluorescent images of parasites at schizont (center) and merozoite stages (outer).
    Figure Legend Snippet: P. falciparum infected RBCs were stained with SYBR Green I and were then examined using a microscope with (a) DAPI and (b) fluorescence filter. Photographs indicated fluorescent images of parasites at schizont (center) and merozoite stages (outer).

    Techniques Used: Infection, Staining, SYBR Green Assay, Microscopy, Fluorescence

    Representative two-channel (FL-1/FL-2) dot-plots, examining various levels of parasitemia. Cultures of P. falciparum were stained with SYBR Green I and CD235A (a–h), and Propidium Iodide and Anti-H (i-j). Over 5,000 events were acquired for each dotplot. (a) Normal unstained RBCs were plotted on the lower left corner. (b) Anti-CD235A PE stained RBCs were plotted between 10 1 and 10 3 on the red axis with small diagonal stretch from the upper left corner (FL2). (c) P. falciparum infected RBC populations were plotted parallel to the noninfected RBC between 10 1 and 10 4 on the green axis (FL1). (d) P. falciparum infected RBC populations controls stained with SYBR Green I and CD235A. Suspected external merozoites from RBCs were localized on the right lower quadrant. Synchronized cultures of P. falciparum with parasitemia were used. (e) 0.01%, (f) 0.5%, (g) 10.0%, and (h) 22.0%.
    Figure Legend Snippet: Representative two-channel (FL-1/FL-2) dot-plots, examining various levels of parasitemia. Cultures of P. falciparum were stained with SYBR Green I and CD235A (a–h), and Propidium Iodide and Anti-H (i-j). Over 5,000 events were acquired for each dotplot. (a) Normal unstained RBCs were plotted on the lower left corner. (b) Anti-CD235A PE stained RBCs were plotted between 10 1 and 10 3 on the red axis with small diagonal stretch from the upper left corner (FL2). (c) P. falciparum infected RBC populations were plotted parallel to the noninfected RBC between 10 1 and 10 4 on the green axis (FL1). (d) P. falciparum infected RBC populations controls stained with SYBR Green I and CD235A. Suspected external merozoites from RBCs were localized on the right lower quadrant. Synchronized cultures of P. falciparum with parasitemia were used. (e) 0.01%, (f) 0.5%, (g) 10.0%, and (h) 22.0%.

    Techniques Used: Staining, SYBR Green Assay, Infection

    21) Product Images from "Gametocytocidal Screen Identifies Novel Chemical Classes with Plasmodium falciparum Transmission Blocking Activity"

    Article Title: Gametocytocidal Screen Identifies Novel Chemical Classes with Plasmodium falciparum Transmission Blocking Activity

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0105817

    Inhibition by MMV Malaria Box. SYBR Green I assay results for the MMV box screened at 10 µM. Plot of percentage of gametocytocidal activity of 400 compounds compared to pyrvinium pamoate control.
    Figure Legend Snippet: Inhibition by MMV Malaria Box. SYBR Green I assay results for the MMV box screened at 10 µM. Plot of percentage of gametocytocidal activity of 400 compounds compared to pyrvinium pamoate control.

    Techniques Used: Inhibition, SYBR Green Assay, Activity Assay

    Overlap of recent screening assays for Malaria Box. Venn Diagram comparing our SYBR Green I assay (green) MMV box hits with hits from four other assays: Confocal fluorescence microscopy (red), Alamar blue early (dark blue) and late (light blue) and Luciferase (yellow) [10] , [14] – [16] .
    Figure Legend Snippet: Overlap of recent screening assays for Malaria Box. Venn Diagram comparing our SYBR Green I assay (green) MMV box hits with hits from four other assays: Confocal fluorescence microscopy (red), Alamar blue early (dark blue) and late (light blue) and Luciferase (yellow) [10] , [14] – [16] .

    Techniques Used: SYBR Green Assay, Fluorescence, Microscopy, Luciferase

    SYBR Green I-Background suppressor gametocytocidal assay. (A) Giemsa stained culture before and after treatment with 10 µM pyrvinium pamoate. (B) SYBR Green I fluorescence of gametocytes, total (diamond), killed (triangle) and live gametocytes after drug treatment (total minus killed, square) with decreasing number of gametocytes per uninfected cell, diluted with 2% hematocrit RBCs in media in presence of background suppressor. (C) Z-factors calculated for each gametocyte dilution. Z factors were calculated using the equation shown, described previously for validating high throughput assays (σ = standard deviation, μ = mean, s = sample, c = control or in this case zero gametocytes) [10] . (D) SYBR Green I fluorescence of live or pyrvinium pamoate-killed gametocytes in the presence of CyQUANT background suppressor, with and without exflagellation with background well fluorescence (no parasites) subtracted out as a blank. (E) Example of assay plate SYBR Green I fluorescence in the presence of background suppressor and calculations for % inhibition. Green indicates a positive hit with high inhibition attributable to gametocyte killing and red indicates an intermediate hit, with inhibition attributable to potentially exflagellation inhibition and/or moderate gametocyte killing. (F) Overall assay setup with five steps: 1. Culture and enrich gametocytes, 2. Incubate with drug for 48 hr, 3. Add exflagellation media and incubate 30 min, 4. Add SYBR Green I and background suppressor and incubate 2 hr, 5. Read SYBR Green I fluorescence at excitation 485 nm and emission 535 nm.
    Figure Legend Snippet: SYBR Green I-Background suppressor gametocytocidal assay. (A) Giemsa stained culture before and after treatment with 10 µM pyrvinium pamoate. (B) SYBR Green I fluorescence of gametocytes, total (diamond), killed (triangle) and live gametocytes after drug treatment (total minus killed, square) with decreasing number of gametocytes per uninfected cell, diluted with 2% hematocrit RBCs in media in presence of background suppressor. (C) Z-factors calculated for each gametocyte dilution. Z factors were calculated using the equation shown, described previously for validating high throughput assays (σ = standard deviation, μ = mean, s = sample, c = control or in this case zero gametocytes) [10] . (D) SYBR Green I fluorescence of live or pyrvinium pamoate-killed gametocytes in the presence of CyQUANT background suppressor, with and without exflagellation with background well fluorescence (no parasites) subtracted out as a blank. (E) Example of assay plate SYBR Green I fluorescence in the presence of background suppressor and calculations for % inhibition. Green indicates a positive hit with high inhibition attributable to gametocyte killing and red indicates an intermediate hit, with inhibition attributable to potentially exflagellation inhibition and/or moderate gametocyte killing. (F) Overall assay setup with five steps: 1. Culture and enrich gametocytes, 2. Incubate with drug for 48 hr, 3. Add exflagellation media and incubate 30 min, 4. Add SYBR Green I and background suppressor and incubate 2 hr, 5. Read SYBR Green I fluorescence at excitation 485 nm and emission 535 nm.

    Techniques Used: SYBR Green Assay, Staining, Fluorescence, High Throughput Screening Assay, Standard Deviation, CyQUANT Assay, Inhibition

    Inhibition by FDA drug library. SYBR Green I assay results for the Johns Hopkins Clinical Compound Library version 1.3 of FDA approved drugs screened at 20 µM. Plot of percentage of gametocytocidal activity of 1,584 compounds compared to clotrimazole control.
    Figure Legend Snippet: Inhibition by FDA drug library. SYBR Green I assay results for the Johns Hopkins Clinical Compound Library version 1.3 of FDA approved drugs screened at 20 µM. Plot of percentage of gametocytocidal activity of 1,584 compounds compared to clotrimazole control.

    Techniques Used: Inhibition, SYBR Green Assay, Activity Assay

    22) Product Images from "A Rapid Growth-Independent Antibiotic Resistance Detection Test by SYBR Green/Propidium Iodide Viability Assay"

    Article Title: A Rapid Growth-Independent Antibiotic Resistance Detection Test by SYBR Green/Propidium Iodide Viability Assay

    Journal: Frontiers in Medicine

    doi: 10.3389/fmed.2018.00127

    SYBR Green I/propidium iodide (PI) assay reveals killing of bactericidal and bacteriostatic antibiotics in 30 min. SYBR Green I/PI assay reveals killing between sensitive and resistant strains after treatment with increasing concentrations of bactericidal drugs (A) kanamycin for Staphylococcus aureus and (B) ampicillin for Escherichia coli . Upon treatment with increasing concentrations of bacteriostatic drugs (C) erythromycin for S. aureus and (D) trimethoprim for E. coli , SYBR Green I/PI assay was consistently able to distinguish sensitive and resistant strains when administered at different concentrations for both sensitive and resistant strains (Student’s t -test, * p
    Figure Legend Snippet: SYBR Green I/propidium iodide (PI) assay reveals killing of bactericidal and bacteriostatic antibiotics in 30 min. SYBR Green I/PI assay reveals killing between sensitive and resistant strains after treatment with increasing concentrations of bactericidal drugs (A) kanamycin for Staphylococcus aureus and (B) ampicillin for Escherichia coli . Upon treatment with increasing concentrations of bacteriostatic drugs (C) erythromycin for S. aureus and (D) trimethoprim for E. coli , SYBR Green I/PI assay was consistently able to distinguish sensitive and resistant strains when administered at different concentrations for both sensitive and resistant strains (Student’s t -test, * p

    Techniques Used: SYBR Green Assay

    SYBR Green I/propidium iodide (PI) stain can distinguish between R (resistant) and S (sensitive) strains of Staphylococcus aureus against various antibiotics in 30 min. Treatment with (A) gentamicin (100 µg/ml), (B) kanamycin (100 µg/ml), (C) erythromycin (400 µg/ml), and (D) ciprofloxacin (100 µg/ml) was added to various overnight S. aureus strains diluted to 1:25 (OD600 = 0.1). After incubation with antibiotics for 30 min, SYBR Green I/PI staining was performed and distinguished the strains and their respective susceptibility categories. All susceptibility results were in concordance with results from the Kirby–Bauer disk diffusion test (Student’s t -test, * p
    Figure Legend Snippet: SYBR Green I/propidium iodide (PI) stain can distinguish between R (resistant) and S (sensitive) strains of Staphylococcus aureus against various antibiotics in 30 min. Treatment with (A) gentamicin (100 µg/ml), (B) kanamycin (100 µg/ml), (C) erythromycin (400 µg/ml), and (D) ciprofloxacin (100 µg/ml) was added to various overnight S. aureus strains diluted to 1:25 (OD600 = 0.1). After incubation with antibiotics for 30 min, SYBR Green I/PI staining was performed and distinguished the strains and their respective susceptibility categories. All susceptibility results were in concordance with results from the Kirby–Bauer disk diffusion test (Student’s t -test, * p

    Techniques Used: SYBR Green Assay, Staining, Incubation, Diffusion-based Assay

    SYBR Green I/propidium iodide (PI) stain can distinguish between R (resistant) and S (sensitive) strains of Gram-negative pathogens against various antibiotics in 30 min. Treatment with (A) ampicillin (100 µg/ml), (B) trimethoprim (50 µg/ml), and (C) streptomycin (50 µg/ml) was added to overnight cultures of Escherichia coli strains diluted to 1:25 (OD600 = 0.1). Treatment with (D) ceftriaxone (25 µg/ml) and (E) cefotaxime (50 µg/ml) was added to overnight cultures of Klebsiella pneumoniae strains diluted to 1:25 (OD600 = 0.1). After incubation with antibiotics for 30 min, SYBR Green I/PI staining was performed and distinguished their respective susceptibility categories. All susceptibility results were in concordance with results from the Kirby–Bauer disk diffusion test (Student’s t -test, * p
    Figure Legend Snippet: SYBR Green I/propidium iodide (PI) stain can distinguish between R (resistant) and S (sensitive) strains of Gram-negative pathogens against various antibiotics in 30 min. Treatment with (A) ampicillin (100 µg/ml), (B) trimethoprim (50 µg/ml), and (C) streptomycin (50 µg/ml) was added to overnight cultures of Escherichia coli strains diluted to 1:25 (OD600 = 0.1). Treatment with (D) ceftriaxone (25 µg/ml) and (E) cefotaxime (50 µg/ml) was added to overnight cultures of Klebsiella pneumoniae strains diluted to 1:25 (OD600 = 0.1). After incubation with antibiotics for 30 min, SYBR Green I/PI staining was performed and distinguished their respective susceptibility categories. All susceptibility results were in concordance with results from the Kirby–Bauer disk diffusion test (Student’s t -test, * p

    Techniques Used: SYBR Green Assay, Staining, Incubation, Diffusion-based Assay

    Drug susceptibility testing of Mycobacterium tuberculosis against first-line tuberculosis drugs using the SYBR Green I/propidium iodide (PI) assay. (A) M. tuberculosis H37Ra and INH-resistant mutants I2, I4 (10-day old) were treated with 10, 500, and 1,000 µg/ml INH overnight (16 h). INH-resistant mutants I2 and I4 were determined to be resistant using 500 and 1,000 µg/ml INH. (B) M. tuberculosis parental strain H37Ra and pyrazinamide (PZA)-resistant mutants (P5 and P2) (20-day old) were treated with PZA (2 mg/ml) overnight (16 h). Treatment with salicylic acid (40 µg/ml) strongly increased the efficacy SYBR Green I/PI assay in detecting PZA resistance in mutants P5 and P2 compared to parental strain H37Ra (Student’s t -test, * p
    Figure Legend Snippet: Drug susceptibility testing of Mycobacterium tuberculosis against first-line tuberculosis drugs using the SYBR Green I/propidium iodide (PI) assay. (A) M. tuberculosis H37Ra and INH-resistant mutants I2, I4 (10-day old) were treated with 10, 500, and 1,000 µg/ml INH overnight (16 h). INH-resistant mutants I2 and I4 were determined to be resistant using 500 and 1,000 µg/ml INH. (B) M. tuberculosis parental strain H37Ra and pyrazinamide (PZA)-resistant mutants (P5 and P2) (20-day old) were treated with PZA (2 mg/ml) overnight (16 h). Treatment with salicylic acid (40 µg/ml) strongly increased the efficacy SYBR Green I/PI assay in detecting PZA resistance in mutants P5 and P2 compared to parental strain H37Ra (Student’s t -test, * p

    Techniques Used: SYBR Green Assay

    A linear relationship between the percentage of live cells and the green/red fluorescence ratio from the SYBR Green I/propidium iodide (PI) viability assay for different bacterial species. Known proportions of isopropyl killed (30 min) and live (A) Staphylococcus aureus (USA300), (B) Klebsiella pneumoniae (Isolate 7), (C) Escherichia coli (W3110), (D) Acinetobacter baumannii , and (E) Mycobacterium tuberculosis (H37Ra) were stained with SYBR Green I/PI and measured using a fluorescence plate reader. A linear regression line was determined. (F) Fluorescence microscopy image showing known proportions (0, 50, and 100%) of live (i–iii) S. aureus (USA300) and (iv–vi) K. pneumoniae (isolate 7) stained with SYBR Green I/PI reveal red (dead) and green (live) ratios in concordance with the known proportions of live and killed organisms from representative Gram-positive ( S. aureus ) and Gram-negative ( K. pneumoniae ) bacteria. Data represent the means ± SEMs.
    Figure Legend Snippet: A linear relationship between the percentage of live cells and the green/red fluorescence ratio from the SYBR Green I/propidium iodide (PI) viability assay for different bacterial species. Known proportions of isopropyl killed (30 min) and live (A) Staphylococcus aureus (USA300), (B) Klebsiella pneumoniae (Isolate 7), (C) Escherichia coli (W3110), (D) Acinetobacter baumannii , and (E) Mycobacterium tuberculosis (H37Ra) were stained with SYBR Green I/PI and measured using a fluorescence plate reader. A linear regression line was determined. (F) Fluorescence microscopy image showing known proportions (0, 50, and 100%) of live (i–iii) S. aureus (USA300) and (iv–vi) K. pneumoniae (isolate 7) stained with SYBR Green I/PI reveal red (dead) and green (live) ratios in concordance with the known proportions of live and killed organisms from representative Gram-positive ( S. aureus ) and Gram-negative ( K. pneumoniae ) bacteria. Data represent the means ± SEMs.

    Techniques Used: Fluorescence, SYBR Green Assay, Viability Assay, Staining, Microscopy

    23) Product Images from "Diversity and quantity of ammonia-oxidizing Archaea and Bacteria in sediment of the Pearl River Estuary, China"

    Article Title: Diversity and quantity of ammonia-oxidizing Archaea and Bacteria in sediment of the Pearl River Estuary, China

    Journal: Applied Microbiology and Biotechnology

    doi: 10.1007/s00253-011-3107-8

    Abundance of AOA amo A, AOB amo A, AOB 16S rRNA genes in the sediments of the Pearl River Estuary. qPCR was performed with three replicates for each sample. For AOA amo A gene, qPCR was conducted using FailSafe™ PCR Premix F and 0.5× SYBR® Green I (Invitrogen, Eugene, USA). For AOB 16S rRNA and amo A genes, PCR was performed using iQ™ SYBR® Green Super Mix. The qPCR thermocycling steps were set as follows: 95°C for 4 min and 45 cycles at 95°C for 45 s, 55°C for 45 s, and 72°C for 45 s. After qPCR assay, the specificity of amplification was verified by melting curve analysis and checking with agarose gel electrophoresis
    Figure Legend Snippet: Abundance of AOA amo A, AOB amo A, AOB 16S rRNA genes in the sediments of the Pearl River Estuary. qPCR was performed with three replicates for each sample. For AOA amo A gene, qPCR was conducted using FailSafe™ PCR Premix F and 0.5× SYBR® Green I (Invitrogen, Eugene, USA). For AOB 16S rRNA and amo A genes, PCR was performed using iQ™ SYBR® Green Super Mix. The qPCR thermocycling steps were set as follows: 95°C for 4 min and 45 cycles at 95°C for 45 s, 55°C for 45 s, and 72°C for 45 s. After qPCR assay, the specificity of amplification was verified by melting curve analysis and checking with agarose gel electrophoresis

    Techniques Used: Real-time Polymerase Chain Reaction, Polymerase Chain Reaction, SYBR Green Assay, Amplification, Agarose Gel Electrophoresis

    24) Product Images from "Dendrimers and Polyamino-Phenolic Ligands: Activity of New Molecules Against Legionella pneumophila Biofilms"

    Article Title: Dendrimers and Polyamino-Phenolic Ligands: Activity of New Molecules Against Legionella pneumophila Biofilms

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2016.00289

    Representative dot plot (FL1 vs. FL3). Planktonic E. coli cells labeled with SYBR Green I and PI treated with G2 dendrimer (B) respect to the untreated control (A) .
    Figure Legend Snippet: Representative dot plot (FL1 vs. FL3). Planktonic E. coli cells labeled with SYBR Green I and PI treated with G2 dendrimer (B) respect to the untreated control (A) .

    Techniques Used: Labeling, SYBR Green Assay

    25) Product Images from "Flow Cytometric Enumeration of Parasitemia in Cultures of Plasmodium falciparum Stained with SYBR Green I and CD235A"

    Article Title: Flow Cytometric Enumeration of Parasitemia in Cultures of Plasmodium falciparum Stained with SYBR Green I and CD235A

    Journal: The Scientific World Journal

    doi: 10.1155/2014/536723

    P. falciparum infected RBCs were stained with SYBR Green I and were then examined using a microscope with (a) DAPI and (b) fluorescence filter. Photographs indicated fluorescent images of parasites at schizont (center) and merozoite stages (outer).
    Figure Legend Snippet: P. falciparum infected RBCs were stained with SYBR Green I and were then examined using a microscope with (a) DAPI and (b) fluorescence filter. Photographs indicated fluorescent images of parasites at schizont (center) and merozoite stages (outer).

    Techniques Used: Infection, Staining, SYBR Green Assay, Microscopy, Fluorescence

    Representative two-channel (FL-1/FL-2) dot-plots, examining various levels of parasitemia. Cultures of P. falciparum were stained with SYBR Green I and CD235A (a–h), and Propidium Iodide and Anti-H (i-j). Over 5,000 events were acquired for each dotplot. (a) Normal unstained RBCs were plotted on the lower left corner. (b) Anti-CD235A PE stained RBCs were plotted between 10 1 and 10 3 on the red axis with small diagonal stretch from the upper left corner (FL2). (c) P. falciparum infected RBC populations were plotted parallel to the noninfected RBC between 10 1 and 10 4 on the green axis (FL1). (d) P. falciparum infected RBC populations controls stained with SYBR Green I and CD235A. Suspected external merozoites from RBCs were localized on the right lower quadrant. Synchronized cultures of P. falciparum with parasitemia were used. (e) 0.01%, (f) 0.5%, (g) 10.0%, and (h) 22.0%.
    Figure Legend Snippet: Representative two-channel (FL-1/FL-2) dot-plots, examining various levels of parasitemia. Cultures of P. falciparum were stained with SYBR Green I and CD235A (a–h), and Propidium Iodide and Anti-H (i-j). Over 5,000 events were acquired for each dotplot. (a) Normal unstained RBCs were plotted on the lower left corner. (b) Anti-CD235A PE stained RBCs were plotted between 10 1 and 10 3 on the red axis with small diagonal stretch from the upper left corner (FL2). (c) P. falciparum infected RBC populations were plotted parallel to the noninfected RBC between 10 1 and 10 4 on the green axis (FL1). (d) P. falciparum infected RBC populations controls stained with SYBR Green I and CD235A. Suspected external merozoites from RBCs were localized on the right lower quadrant. Synchronized cultures of P. falciparum with parasitemia were used. (e) 0.01%, (f) 0.5%, (g) 10.0%, and (h) 22.0%.

    Techniques Used: Staining, SYBR Green Assay, Infection

    26) Product Images from "Alteration of Sexual Reproduction and Genetic Diversity in the Kelp Species Laminaria digitata at the Southern Limit of Its Range"

    Article Title: Alteration of Sexual Reproduction and Genetic Diversity in the Kelp Species Laminaria digitata at the Southern Limit of Its Range

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0102518

    Example flow cytometric analysis of spore ploidy, showing histograms of Sybr Green I fluorescence (DNA content). Grey lines show a representative Quiberon sporophyte and black lines show a representative Porspoder sporophyte. In both cases the thick lines represent spores, and thin lines represent the internal standard ( Emiliania huxleyi cells) added to the same sample during staining.
    Figure Legend Snippet: Example flow cytometric analysis of spore ploidy, showing histograms of Sybr Green I fluorescence (DNA content). Grey lines show a representative Quiberon sporophyte and black lines show a representative Porspoder sporophyte. In both cases the thick lines represent spores, and thin lines represent the internal standard ( Emiliania huxleyi cells) added to the same sample during staining.

    Techniques Used: Flow Cytometry, SYBR Green Assay, Fluorescence, Staining

    27) Product Images from "Targeted disruption of the Chop gene delays endoplasmic reticulum stress-mediated diabetes"

    Article Title: Targeted disruption of the Chop gene delays endoplasmic reticulum stress-mediated diabetes

    Journal: The Journal of Clinical Investigation

    doi: 10.1172/JCI14550

    Apoptosis in MIN6 cells overexpressing Ins2 C96Y –EGFP and Ins2 C96Y . ( a ) Cells were transfected with p Ins2 WT -EGFP or p Ins2 C96Y -EGFP. At the indicated times after transfection, cells were observed under a fluorescence microscope. Original magnification: ×100. ( b ) Cells were cotransfected with pEYFP-ER and either pcDNA- Ins2 WT or pcDNA- Ins2 C96Y . Forty-eight hours after transfection, cells were observed under a fluorescence microscope. Original magnification: ×200. ( c ) Cells were cotransfected with pEGFP and either pcDNA- Ins2 WT or pcDNA- Ins2 C96Y . The transfected cells and apoptotic cells were visualized by GFP fluorescence and Hoechst 33258 staining, respectively. pcDNA- Ins2 WT –transfected cells were not apoptotic (arrowheads), whereas pcDNA- Ins2 C96Y –transfected cells were apoptotic (arrows). Original magnification: ×400. ( d ) Cells were transfected with pcDNA- Ins2 WT or pcDNA- Ins2 C96Y . DNA was extracted, electrophoresed in 2% agarose gel, stained with SYBR Green I, and visualized by UV transillumination. ( e ) Cells were cotransfected with pEGFP and either pcDNA- Ins2 WT or pcDNA- Ins2 C96Y . The transfected cells and apoptotic cells were visualized by GFP fluorescence and annexin V staining, respectively. Original magnification: ×200.
    Figure Legend Snippet: Apoptosis in MIN6 cells overexpressing Ins2 C96Y –EGFP and Ins2 C96Y . ( a ) Cells were transfected with p Ins2 WT -EGFP or p Ins2 C96Y -EGFP. At the indicated times after transfection, cells were observed under a fluorescence microscope. Original magnification: ×100. ( b ) Cells were cotransfected with pEYFP-ER and either pcDNA- Ins2 WT or pcDNA- Ins2 C96Y . Forty-eight hours after transfection, cells were observed under a fluorescence microscope. Original magnification: ×200. ( c ) Cells were cotransfected with pEGFP and either pcDNA- Ins2 WT or pcDNA- Ins2 C96Y . The transfected cells and apoptotic cells were visualized by GFP fluorescence and Hoechst 33258 staining, respectively. pcDNA- Ins2 WT –transfected cells were not apoptotic (arrowheads), whereas pcDNA- Ins2 C96Y –transfected cells were apoptotic (arrows). Original magnification: ×400. ( d ) Cells were transfected with pcDNA- Ins2 WT or pcDNA- Ins2 C96Y . DNA was extracted, electrophoresed in 2% agarose gel, stained with SYBR Green I, and visualized by UV transillumination. ( e ) Cells were cotransfected with pEGFP and either pcDNA- Ins2 WT or pcDNA- Ins2 C96Y . The transfected cells and apoptotic cells were visualized by GFP fluorescence and annexin V staining, respectively. Original magnification: ×200.

    Techniques Used: Transfection, Fluorescence, Microscopy, Staining, Agarose Gel Electrophoresis, SYBR Green Assay

    28) Product Images from "Development of a Real-Time PCR Assay Using SYBR Green I for Provirus Load Quantification in a Murine Model of AIDS"

    Article Title: Development of a Real-Time PCR Assay Using SYBR Green I for Provirus Load Quantification in a Murine Model of AIDS

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.42.9.4361-4364.2004

    BM5d provirus load in infected (•) and AZT-DDI-treated (○) mice, as measured by real-time SYBR Green I PCR, in tissues and macrophages as described in the text. The BM5d copies were deduced from linear regression analyses of the standard curve and divided by the actual number of cells, calculated after 18S rRNA gene normalization, to obtain the BM5d provirus copies/cell. Bars show the mean for each group. The real-time PCR performed on mock-infected (control) mice gave negative results.
    Figure Legend Snippet: BM5d provirus load in infected (•) and AZT-DDI-treated (○) mice, as measured by real-time SYBR Green I PCR, in tissues and macrophages as described in the text. The BM5d copies were deduced from linear regression analyses of the standard curve and divided by the actual number of cells, calculated after 18S rRNA gene normalization, to obtain the BM5d provirus copies/cell. Bars show the mean for each group. The real-time PCR performed on mock-infected (control) mice gave negative results.

    Techniques Used: Infection, Mouse Assay, SYBR Green Assay, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction

    29) Product Images from "Extracellular DNA release confers heterogeneity in Candida albicans biofilm formation"

    Article Title: Extracellular DNA release confers heterogeneity in Candida albicans biofilm formation

    Journal: BMC Microbiology

    doi: 10.1186/s12866-014-0303-6

    Variation in C. albicans biofilm formation and eDNA release. C. albicans isolates with LBF (n = 3) and HBF (n = 3) were grown as biofilms in 96 well flat-bottom microtitre plates at 37°C for 24 h. (A) Biofilm biomass was assessed spectrophotometrically by reading CV absorbance, data represents mean ± SD. (B) Biofilms were treated ± 256 μg/ml DNase for a further 24 h before being passed through 0.22 μM membrane filter. Biomass retained on the filters was dried overnight at 60°C and dry weight measurements taken. In addition, biofilms were stained with CV and imaged to show the disruptive effect of DNase on the biofilms, data represents mean ± SE. (C) Isolates were grown as biofilms for 4 and 24 h in the presence of the DNA binding dye SYBR® Green I. Fluorescence was measured after 4 and 24 h at Ex485/Em518. Absorbance was measured simultaneously for normalising the fluorescence data, data represents mean ± SD. Each isolate was tested in duplicate, on three independent occasions. * # p
    Figure Legend Snippet: Variation in C. albicans biofilm formation and eDNA release. C. albicans isolates with LBF (n = 3) and HBF (n = 3) were grown as biofilms in 96 well flat-bottom microtitre plates at 37°C for 24 h. (A) Biofilm biomass was assessed spectrophotometrically by reading CV absorbance, data represents mean ± SD. (B) Biofilms were treated ± 256 μg/ml DNase for a further 24 h before being passed through 0.22 μM membrane filter. Biomass retained on the filters was dried overnight at 60°C and dry weight measurements taken. In addition, biofilms were stained with CV and imaged to show the disruptive effect of DNase on the biofilms, data represents mean ± SE. (C) Isolates were grown as biofilms for 4 and 24 h in the presence of the DNA binding dye SYBR® Green I. Fluorescence was measured after 4 and 24 h at Ex485/Em518. Absorbance was measured simultaneously for normalising the fluorescence data, data represents mean ± SD. Each isolate was tested in duplicate, on three independent occasions. * # p

    Techniques Used: Staining, Binding Assay, SYBR Green Assay, Fluorescence

    30) Product Images from "Nutrient requirements for growth of the extreme oligotroph 'Candidatus Pelagibacter ubique' HTCC1062 on a defined medium"

    Article Title: Nutrient requirements for growth of the extreme oligotroph 'Candidatus Pelagibacter ubique' HTCC1062 on a defined medium

    Journal: The ISME Journal

    doi: 10.1038/ismej.2012.122

    DNA content and morphology of SYBR Green I-stained stationary-phase cells from pyruvate-deplete and -replete batch cultures. Red dashed line in a and b represents the minimum threshold of fluorescence detection. Black dashed lines in a and b represent relative DNA fluorescence values of 300–325 per event and 475–500 per event, as indicated with black arrows. ( a ) Relative DNA fluorescence of cells from pyruvate-replete (50 μℳ) stationary-phase cultures, and ( b ) pyruvate-deplete (0.5 μℳ) stationary-phase cultures. ( c ) Fluorescent microscopy image of cells from ( a ). Arrowheads point to single cells. ( d ) Fluorescent microscopy image of cells from ( b ). Arrowheads point to cell doublets.
    Figure Legend Snippet: DNA content and morphology of SYBR Green I-stained stationary-phase cells from pyruvate-deplete and -replete batch cultures. Red dashed line in a and b represents the minimum threshold of fluorescence detection. Black dashed lines in a and b represent relative DNA fluorescence values of 300–325 per event and 475–500 per event, as indicated with black arrows. ( a ) Relative DNA fluorescence of cells from pyruvate-replete (50 μℳ) stationary-phase cultures, and ( b ) pyruvate-deplete (0.5 μℳ) stationary-phase cultures. ( c ) Fluorescent microscopy image of cells from ( a ). Arrowheads point to single cells. ( d ) Fluorescent microscopy image of cells from ( b ). Arrowheads point to cell doublets.

    Techniques Used: SYBR Green Assay, Staining, Fluorescence, Microscopy

    31) Product Images from "Identification of pork in meat products using real-time loop-mediated isothermal amplification"

    Article Title: Identification of pork in meat products using real-time loop-mediated isothermal amplification

    Journal: Biotechnology, Biotechnological Equipment

    doi: 10.1080/13102818.2014.963789

    SYBR green I fluorescent dye-mediated monitoring of pork specific RealAmp assay amplification.
    Figure Legend Snippet: SYBR green I fluorescent dye-mediated monitoring of pork specific RealAmp assay amplification.

    Techniques Used: SYBR Green Assay, Amplification

    32) Product Images from "A Comparison of Methods for Counting Viruses in Aquatic Systems"

    Article Title: A Comparison of Methods for Counting Viruses in Aquatic Systems

    Journal: Applied and Environmental Microbiology

    doi:

    EM-SYBR Green I protocol: relationship between the mean viral concentrations and the corresponding coefficients of variation (CV) for the six environments tested.
    Figure Legend Snippet: EM-SYBR Green I protocol: relationship between the mean viral concentrations and the corresponding coefficients of variation (CV) for the six environments tested.

    Techniques Used: SYBR Green Assay

    33) Product Images from "Nanocalorimetric Characterization of Microbial Activity in Deep Subsurface Oceanic Crustal Fluids"

    Article Title: Nanocalorimetric Characterization of Microbial Activity in Deep Subsurface Oceanic Crustal Fluids

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2016.00454

    Epifluorescence-microscopy image of basaltic crustal fluid samples from JFR filtered onto a Whatman 0.2 μm Anodisc filter stained with SYBR Green I, before (A) and after (B) induction of spore germination by pasteurization and 48 h of incubation at 50°C. Scale bar 10 μm.
    Figure Legend Snippet: Epifluorescence-microscopy image of basaltic crustal fluid samples from JFR filtered onto a Whatman 0.2 μm Anodisc filter stained with SYBR Green I, before (A) and after (B) induction of spore germination by pasteurization and 48 h of incubation at 50°C. Scale bar 10 μm.

    Techniques Used: Epifluorescence Microscopy, Staining, SYBR Green Assay, Incubation

    34) Product Images from "Loop-mediated Isothermal Amplification assay for Detection of Candidatus Liberibacter Asiaticus, a Causal Agent of Citrus Huanglongbing"

    Article Title: Loop-mediated Isothermal Amplification assay for Detection of Candidatus Liberibacter Asiaticus, a Causal Agent of Citrus Huanglongbing

    Journal: The Plant Pathology Journal

    doi: 10.5423/PPJ.FT.10.2018.0212

    Loop-mediated isothermal amplification (LAMP) for specific detection of ‘ Candidatus Liberibacter asiaticus’ using the primer set from the prophage gene, LasA I in HLB-infected leaves of grapefruit according to reaction temperatures (57, 60, 62 and 65°C). condition test for HLB detection. (A) Visual detection under normal light by adding SYBR Green I dye. (B) Electrophoresis analysis on 1% agarose gel. Lanes 1–4; HLB-infected grapefruit leaves, lanes 5–8; healthy grapefruit leaves, lanes 9–11; distilled water, lane M; 100 bp DNA ladder (NEB New England Biolabs, cat# N3231S).
    Figure Legend Snippet: Loop-mediated isothermal amplification (LAMP) for specific detection of ‘ Candidatus Liberibacter asiaticus’ using the primer set from the prophage gene, LasA I in HLB-infected leaves of grapefruit according to reaction temperatures (57, 60, 62 and 65°C). condition test for HLB detection. (A) Visual detection under normal light by adding SYBR Green I dye. (B) Electrophoresis analysis on 1% agarose gel. Lanes 1–4; HLB-infected grapefruit leaves, lanes 5–8; healthy grapefruit leaves, lanes 9–11; distilled water, lane M; 100 bp DNA ladder (NEB New England Biolabs, cat# N3231S).

    Techniques Used: Amplification, Infection, SYBR Green Assay, Electrophoresis, Agarose Gel Electrophoresis

    35) Product Images from "Optimization of loop-mediated isothermal amplification (LAMP) assays for the detection of Leishmania DNA in human blood samples"

    Article Title: Optimization of loop-mediated isothermal amplification (LAMP) assays for the detection of Leishmania DNA in human blood samples

    Journal: Acta Tropica

    doi: 10.1016/j.actatropica.2016.06.009

    Agarose gel electrophoresis analysis and SYBR Green I end point detection of LITS-LAMP1 (A,B), and LITS-LAMP2 (C,D). The analysis shows LAMP DNA amplification products of different concentrations amplified from different amounts of L. donovani template DNA, (1) 0.1 ng, (2) 0.01 ng, (3) 1 pg, (4) 0.1 pg. And the amplification of 1 ng template DNA of each of the following Leishmania species: (5) L. major , (6) L. aethiopica , (7) L. tropica , (8) L. infantum chagasi . (9) and (10) No DNA [control], (11–16) Select blood samples obtained by finger prick from volunteers in North Ethiopia. (M) DNA size marker.
    Figure Legend Snippet: Agarose gel electrophoresis analysis and SYBR Green I end point detection of LITS-LAMP1 (A,B), and LITS-LAMP2 (C,D). The analysis shows LAMP DNA amplification products of different concentrations amplified from different amounts of L. donovani template DNA, (1) 0.1 ng, (2) 0.01 ng, (3) 1 pg, (4) 0.1 pg. And the amplification of 1 ng template DNA of each of the following Leishmania species: (5) L. major , (6) L. aethiopica , (7) L. tropica , (8) L. infantum chagasi . (9) and (10) No DNA [control], (11–16) Select blood samples obtained by finger prick from volunteers in North Ethiopia. (M) DNA size marker.

    Techniques Used: Agarose Gel Electrophoresis, SYBR Green Assay, Amplification, Marker

    36) Product Images from "Reverse Transcription Loop-Mediated Isothermal Amplification for Rapid Detection of Japanese Encephalitis Virus in Swine and Mosquitoes"

    Article Title: Reverse Transcription Loop-Mediated Isothermal Amplification for Rapid Detection of Japanese Encephalitis Virus in Swine and Mosquitoes

    Journal: Vector Borne and Zoonotic Diseases

    doi: 10.1089/vbz.2012.0991

    Sensitivity of the RT-LAMP assay for the JEV I and JEV III E genes, as detected by electrophoresis and SYBR Green I staining. Shown are the RT-LAMP 2% agarose gel electrophoresis detection limits for JEV I ( A ) and JEV III ( B ), which were 2.57 and 2.34
    Figure Legend Snippet: Sensitivity of the RT-LAMP assay for the JEV I and JEV III E genes, as detected by electrophoresis and SYBR Green I staining. Shown are the RT-LAMP 2% agarose gel electrophoresis detection limits for JEV I ( A ) and JEV III ( B ), which were 2.57 and 2.34

    Techniques Used: RT Lamp Assay, Electrophoresis, SYBR Green Assay, Staining, Agarose Gel Electrophoresis

    37) Product Images from "The Response of Heterotrophic Prokaryote and Viral Communities to Labile Organic Carbon Inputs Is Controlled by the Predator Food Chain Structure"

    Article Title: The Response of Heterotrophic Prokaryote and Viral Communities to Labile Organic Carbon Inputs Is Controlled by the Predator Food Chain Structure

    Journal: Viruses

    doi: 10.3390/v9090238

    Biparametric flow cytrometry virus plots. Four different viral populations were discriminated combining side scatter signal vs. green fluorescent signal after staining with SYBR Green I (reflecting the amount of DNA and hence genome sizes): I, Small viruses, II, medium viruses, III, large viruses, IV, huge viruses ( A ) (see also Figure S2 ). Abundance of small (I), medium (II), large (III), and huge (IV) viruses determined by FCM in PAME-I ( B ). Blue circles show no addition of carbon in the form of glucose, while red circles show three Redfield additions of carbon. Open and filled circles display perturbations with or without silicate (open: no Si addition; filled: Si addition). See Figure 2 for more information about experimental design.
    Figure Legend Snippet: Biparametric flow cytrometry virus plots. Four different viral populations were discriminated combining side scatter signal vs. green fluorescent signal after staining with SYBR Green I (reflecting the amount of DNA and hence genome sizes): I, Small viruses, II, medium viruses, III, large viruses, IV, huge viruses ( A ) (see also Figure S2 ). Abundance of small (I), medium (II), large (III), and huge (IV) viruses determined by FCM in PAME-I ( B ). Blue circles show no addition of carbon in the form of glucose, while red circles show three Redfield additions of carbon. Open and filled circles display perturbations with or without silicate (open: no Si addition; filled: Si addition). See Figure 2 for more information about experimental design.

    Techniques Used: Flow Cytometry, Staining, SYBR Green Assay

    38) Product Images from "Exclusion of small terminase mediated DNA threading models for genome packaging in bacteriophage T4"

    Article Title: Exclusion of small terminase mediated DNA threading models for genome packaging in bacteriophage T4

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkw184

    The oligomerization domain alone is insufficient for DNA binding. ( A ) Schematic of the oligomerization domain deletion construct. Numbers represent the amino acid positions in the gp16 coding sequence. ( B ) Elution profile of the gp16 oligomerization domain deletion mutant. The void volume is indicated by an arrow. Oligomeric state of the domain was calculated from the elution volume. ( C ) The peak fraction from (B) was electrophoresed on a 4–20% gradient native polyacrylamide gel and the gel was stained with Coomassie blue for protein (lane 1) and SYBR Green I for DNA (lane 2). ( D ) In vitro DNA binding of the oligomerization domain deletion mutant. The purified protein (25–75 μM) was incubated with the 500-bp g16 DNA (5 nM) in a 12 μl reaction mixture for 15 min. Samples were then electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with SYBR Green I. The control lane 1 contained no gp16.
    Figure Legend Snippet: The oligomerization domain alone is insufficient for DNA binding. ( A ) Schematic of the oligomerization domain deletion construct. Numbers represent the amino acid positions in the gp16 coding sequence. ( B ) Elution profile of the gp16 oligomerization domain deletion mutant. The void volume is indicated by an arrow. Oligomeric state of the domain was calculated from the elution volume. ( C ) The peak fraction from (B) was electrophoresed on a 4–20% gradient native polyacrylamide gel and the gel was stained with Coomassie blue for protein (lane 1) and SYBR Green I for DNA (lane 2). ( D ) In vitro DNA binding of the oligomerization domain deletion mutant. The purified protein (25–75 μM) was incubated with the 500-bp g16 DNA (5 nM) in a 12 μl reaction mixture for 15 min. Samples were then electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with SYBR Green I. The control lane 1 contained no gp16.

    Techniques Used: Binding Assay, Construct, Sequencing, Mutagenesis, Staining, SYBR Green Assay, In Vitro, Purification, Incubation

    gp16 binds DNA in vitro . The purified phage T4 gp16 (25–75 μM) was incubated with the substrate DNA (500-bp amplified g16 DNA) (5 nM) in a 12 μl reaction mixture for 15 min. The reaction mixture was electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with Coomassie blue for protein ( A ) and SYBR Green I for DNA ( B ). Control lanes were missing either the gp16 protein (lane 1) or the DNA (lane 5). Position of the shifted DNA band is indicated by a dashed line.
    Figure Legend Snippet: gp16 binds DNA in vitro . The purified phage T4 gp16 (25–75 μM) was incubated with the substrate DNA (500-bp amplified g16 DNA) (5 nM) in a 12 μl reaction mixture for 15 min. The reaction mixture was electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with Coomassie blue for protein ( A ) and SYBR Green I for DNA ( B ). Control lanes were missing either the gp16 protein (lane 1) or the DNA (lane 5). Position of the shifted DNA band is indicated by a dashed line.

    Techniques Used: In Vitro, Purification, Incubation, Amplification, Staining, SYBR Green Assay

    Analysis of in vivo -bound gp16-DNA complex. (A and B) Purified gp16 from phage T4 and related phages (8 μg each) were electrophoresed on a native 4–20% gradient polyacrylamide gel. The same gel was stained with Coomassie blue for protein ( A ) and SYBR Green I for DNA ( B ). The arrow corresponds to a fragment of DNA that was dissociated from RB49 gp16 during electrophoresis (panel B, lane 3). (C and D) Most of the gp16-bound DNA is resistant to Benzonase. The purified T4 and RB49 gp16 proteins (1 mg) were treated with Benzonase (Novagen) overnight at room temperature to digest the loosely-bound DNA fragments. Benzonase was removed by passing the samples through a HiLoad 16/600 Superdex 200 size-exclusion column. The tightly bound nuclease-protected DNA was then released by digesting gp16 with Proteinase K (Thermo Scientific) at 65°C for 30 min. Samples were analyzed on a 4–20% gradient polyacrylamide gel. The gel was stained with Coomassie blue for protein ( C ) and SYBR Green I for DNA ( D ). The positions of the DNA bands released from Proteinase K digestion are marked with arrows (panel D, lanes 4 and 7).
    Figure Legend Snippet: Analysis of in vivo -bound gp16-DNA complex. (A and B) Purified gp16 from phage T4 and related phages (8 μg each) were electrophoresed on a native 4–20% gradient polyacrylamide gel. The same gel was stained with Coomassie blue for protein ( A ) and SYBR Green I for DNA ( B ). The arrow corresponds to a fragment of DNA that was dissociated from RB49 gp16 during electrophoresis (panel B, lane 3). (C and D) Most of the gp16-bound DNA is resistant to Benzonase. The purified T4 and RB49 gp16 proteins (1 mg) were treated with Benzonase (Novagen) overnight at room temperature to digest the loosely-bound DNA fragments. Benzonase was removed by passing the samples through a HiLoad 16/600 Superdex 200 size-exclusion column. The tightly bound nuclease-protected DNA was then released by digesting gp16 with Proteinase K (Thermo Scientific) at 65°C for 30 min. Samples were analyzed on a 4–20% gradient polyacrylamide gel. The gel was stained with Coomassie blue for protein ( C ) and SYBR Green I for DNA ( D ). The positions of the DNA bands released from Proteinase K digestion are marked with arrows (panel D, lanes 4 and 7).

    Techniques Used: In Vivo, Purification, Staining, SYBR Green Assay, Electrophoresis

    Deletion of channel helix α2 did not disrupt DNA binding. ( A ) Schematic of the central domain helix α2 deletion mutants. Numbers represent the amino acid positions in the gp16 coding sequence. Deleted sequences are shown by polylines and the amino acid residues flanking the deletions are shown in blue. ( B ) Elution profiles of mutants Del-1 (magenta), Del-2 (blue) and Del-3 (green). The void volume is indicated by an arrow. The oligomeric state of the gp16 species in peak b of the mutants was calculated from the elution volume. (C and D) The peak gp16 fractions from B were electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with Coomassie blue for protein ( C ) and SYBR Green I for DNA ( D ). ‘a’ and ‘b’ correspond to the fractions from the peaks a and b of the respective mutant. ( E ) In vitro DNA binding of the deletion mutants. The purified proteins (20–60 μM) were incubated with the 500-bp g16 DNA (5 nM) in 12 μl reaction mixture for 15 min. Samples were then electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with SYBR Green I. The control lane 1 contained no gp16 and shifted bands are shown by arrows in lanes 12–16.
    Figure Legend Snippet: Deletion of channel helix α2 did not disrupt DNA binding. ( A ) Schematic of the central domain helix α2 deletion mutants. Numbers represent the amino acid positions in the gp16 coding sequence. Deleted sequences are shown by polylines and the amino acid residues flanking the deletions are shown in blue. ( B ) Elution profiles of mutants Del-1 (magenta), Del-2 (blue) and Del-3 (green). The void volume is indicated by an arrow. The oligomeric state of the gp16 species in peak b of the mutants was calculated from the elution volume. (C and D) The peak gp16 fractions from B were electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with Coomassie blue for protein ( C ) and SYBR Green I for DNA ( D ). ‘a’ and ‘b’ correspond to the fractions from the peaks a and b of the respective mutant. ( E ) In vitro DNA binding of the deletion mutants. The purified proteins (20–60 μM) were incubated with the 500-bp g16 DNA (5 nM) in 12 μl reaction mixture for 15 min. Samples were then electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with SYBR Green I. The control lane 1 contained no gp16 and shifted bands are shown by arrows in lanes 12–16.

    Techniques Used: Binding Assay, Sequencing, Staining, SYBR Green Assay, Mutagenesis, In Vitro, Purification, Incubation

    The positively charged residues of gp16 channel are not essential for DNA binding. ( A ) Schematic of the mutations introduced into the helix α2 of T4 gp16. Numbers represent the amino acid positions in the gp16 coding sequence. Positions of the mutated residues are indicated as black bars in helix α2. ( B ) Elution profiles of WT gp16 (black) and mutants 3M (blue) and 4M (red). The void volume is indicated by an arrow. The hexamer size of gp16 species in peak b of mutant 4M was calculated from the elution volume using the column calibrated with molecular weight standards. (C and D) The peak gp16 fractions from B were electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with Coomassie blue for protein ( C ) and SYBR Green I for DNA ( D ). Lanes 4M-a and 4M-b represent the fractions from peak a and peak b of the mutant 4M. ( E ) In vitro DNA binding of gp16 mutants 3M and 4M. Proteins (20–60 μM) were incubated with the 500-bp g16 DNA (5 nM) in 12 μl reaction mixture for 15 min. Samples were then electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with SYBR Green I. The control lane 1 contained no gp16.
    Figure Legend Snippet: The positively charged residues of gp16 channel are not essential for DNA binding. ( A ) Schematic of the mutations introduced into the helix α2 of T4 gp16. Numbers represent the amino acid positions in the gp16 coding sequence. Positions of the mutated residues are indicated as black bars in helix α2. ( B ) Elution profiles of WT gp16 (black) and mutants 3M (blue) and 4M (red). The void volume is indicated by an arrow. The hexamer size of gp16 species in peak b of mutant 4M was calculated from the elution volume using the column calibrated with molecular weight standards. (C and D) The peak gp16 fractions from B were electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with Coomassie blue for protein ( C ) and SYBR Green I for DNA ( D ). Lanes 4M-a and 4M-b represent the fractions from peak a and peak b of the mutant 4M. ( E ) In vitro DNA binding of gp16 mutants 3M and 4M. Proteins (20–60 μM) were incubated with the 500-bp g16 DNA (5 nM) in 12 μl reaction mixture for 15 min. Samples were then electrophoresed on a 4–20% gradient native polyacrylamide gel and stained with SYBR Green I. The control lane 1 contained no gp16.

    Techniques Used: Binding Assay, Sequencing, Mutagenesis, Molecular Weight, Staining, SYBR Green Assay, In Vitro, Incubation

    39) Product Images from "Live Cell Imaging of Butterfly Pupal and Larval Wings In Vivo"

    Article Title: Live Cell Imaging of Butterfly Pupal and Larval Wings In Vivo

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0128332

    Staining pattern of a prospective eyespot area and edge spots. ( a ) A larval wing imaginal disk. Stained with SYBR Green I for nuclei and MitoTracker Orange for mitochondria. A white arrow indicates a prospective eyespot area, as judged by venation pattern. ( b ) Pupal wing tissue stained with SYBR Green I for nuclei and MitoTracker Orange for mitochondria. A white arrow indicates a prospective eyespot area. ( c ) High magnification of a prospective eyespot area shown in (b). ( d ) Peripheral area of a pupal wing tissue stained with CFSE. Arrowheads indicate pupal edge spots.
    Figure Legend Snippet: Staining pattern of a prospective eyespot area and edge spots. ( a ) A larval wing imaginal disk. Stained with SYBR Green I for nuclei and MitoTracker Orange for mitochondria. A white arrow indicates a prospective eyespot area, as judged by venation pattern. ( b ) Pupal wing tissue stained with SYBR Green I for nuclei and MitoTracker Orange for mitochondria. A white arrow indicates a prospective eyespot area. ( c ) High magnification of a prospective eyespot area shown in (b). ( d ) Peripheral area of a pupal wing tissue stained with CFSE. Arrowheads indicate pupal edge spots.

    Techniques Used: Staining, SYBR Green Assay

    Double staining of a larval wing imaginal disk and pupal wing tissue. The tissue was stained with SYBR Green I for nuclei and MitoTacker Orange for mitochondria. Scale bars indicate 10 μm. ( a ) Larval wing imaginal disk. A portion is enlarged in the right panel. An optical vertical cross-section image shown at the bottom was made along a blue broken line. Thickness of a cell was approximately 2.9 μm. The image was constructed by stacking 28 images down to 5.5 μm in depth with 0.2 μm intervals. ( b ) Pupal wing tissue. The pupal head is positioned to the left. A portion is enlarged in the right panel. An optical vertical cross-section image shown at the bottom was made along a blue broken line. Thickness of a cell was more than 11.4 μm (the deepest portion not detected). The image was constructed by stacking 58 images with 0.2 μm intervals.
    Figure Legend Snippet: Double staining of a larval wing imaginal disk and pupal wing tissue. The tissue was stained with SYBR Green I for nuclei and MitoTacker Orange for mitochondria. Scale bars indicate 10 μm. ( a ) Larval wing imaginal disk. A portion is enlarged in the right panel. An optical vertical cross-section image shown at the bottom was made along a blue broken line. Thickness of a cell was approximately 2.9 μm. The image was constructed by stacking 28 images down to 5.5 μm in depth with 0.2 μm intervals. ( b ) Pupal wing tissue. The pupal head is positioned to the left. A portion is enlarged in the right panel. An optical vertical cross-section image shown at the bottom was made along a blue broken line. Thickness of a cell was more than 11.4 μm (the deepest portion not detected). The image was constructed by stacking 58 images with 0.2 μm intervals.

    Techniques Used: Double Staining, Staining, SYBR Green Assay, Construct

    40) Product Images from "Preparation of DNA and protein micro arrays on glass slides coated with an agarose film"

    Article Title: Preparation of DNA and protein micro arrays on glass slides coated with an agarose film

    Journal: Nucleic Acids Research

    doi:

    Match and mismatch discrimination. ( A ) A fluorescence labeled oligonucleotide was hybridized against an array containing oligonucleotides with perfect match (row 1), oligonucleotides with two internal mismatches (row 2) or oligonucleotides with an unrelated sequence (row 3). ( B ) Signal of SYBR Green I staining of the same array after hybridization demonstrating the presence of oligonucleotide probes in all rows. The higher fluorescence signal in the two hybridized rows is probably due to better staining of double-stranded DNA. Both signals are close to saturation. ( C ) Densitometric evaluation of signals as an average of five spots. The distance between the spots was 200 m.
    Figure Legend Snippet: Match and mismatch discrimination. ( A ) A fluorescence labeled oligonucleotide was hybridized against an array containing oligonucleotides with perfect match (row 1), oligonucleotides with two internal mismatches (row 2) or oligonucleotides with an unrelated sequence (row 3). ( B ) Signal of SYBR Green I staining of the same array after hybridization demonstrating the presence of oligonucleotide probes in all rows. The higher fluorescence signal in the two hybridized rows is probably due to better staining of double-stranded DNA. Both signals are close to saturation. ( C ) Densitometric evaluation of signals as an average of five spots. The distance between the spots was 200 m.

    Techniques Used: Fluorescence, Labeling, Sequencing, SYBR Green Assay, Staining, Hybridization

    41) Product Images from "Localization of retinaldehyde dehydrogenases and retinoid binding proteins to sustentacular cells, glia, Bowman's gland cells and stroma: potential sites of retinoic acid synthesis in the postnatal rat olfactory organ"

    Article Title: Localization of retinaldehyde dehydrogenases and retinoid binding proteins to sustentacular cells, glia, Bowman's gland cells and stroma: potential sites of retinoic acid synthesis in the postnatal rat olfactory organ

    Journal: The Journal of comparative neurology

    doi: 10.1002/cne.20904

    PCR reactions with primers specific for RALDH 1, 2, and 3 and ALDH-PB were carried out as described in Materials and Methods using either 100 ng postnatal rat olfactory cDNA ( A) or 100 ng genomic DNA ( B ). A. mRNAs encoding RALDH 1, 2, and 3 are expressed in postnatal rat olfactory tissue. Shown is an image of the gel-separated, RT-PCR products from reactions containing cDNA prepared from DNAse-I treated olfactory tissue RNA. Bands were visualized with SYBR Green I. Lane 1, 100 bp ladder; lane 2, RALDH 1; lane 3, ALDH-PB; lane 4, RALDH 2; lane 5, RALDH 3; lane 6, minus cDNA control reaction containing primers for all ALDH-PB / RALDH transcripts. B. ALDH-PB mRNA was not detected in olfactory tissue, but the ALDH-PB primers amplify authentic gene products from rat genomic DNA. Shown is an image of gel-separated, PCR products from reactions containing rat genomic DNA. Bands were visualized with SYBR Green I. Lane 1, RALDH 1; lane 2, ALDH-PB. Markers designating migration points of 300 and 400 bp ladder transcripts that were run on the gel are shown to the left of the image.
    Figure Legend Snippet: PCR reactions with primers specific for RALDH 1, 2, and 3 and ALDH-PB were carried out as described in Materials and Methods using either 100 ng postnatal rat olfactory cDNA ( A) or 100 ng genomic DNA ( B ). A. mRNAs encoding RALDH 1, 2, and 3 are expressed in postnatal rat olfactory tissue. Shown is an image of the gel-separated, RT-PCR products from reactions containing cDNA prepared from DNAse-I treated olfactory tissue RNA. Bands were visualized with SYBR Green I. Lane 1, 100 bp ladder; lane 2, RALDH 1; lane 3, ALDH-PB; lane 4, RALDH 2; lane 5, RALDH 3; lane 6, minus cDNA control reaction containing primers for all ALDH-PB / RALDH transcripts. B. ALDH-PB mRNA was not detected in olfactory tissue, but the ALDH-PB primers amplify authentic gene products from rat genomic DNA. Shown is an image of gel-separated, PCR products from reactions containing rat genomic DNA. Bands were visualized with SYBR Green I. Lane 1, RALDH 1; lane 2, ALDH-PB. Markers designating migration points of 300 and 400 bp ladder transcripts that were run on the gel are shown to the left of the image.

    Techniques Used: Polymerase Chain Reaction, Reverse Transcription Polymerase Chain Reaction, SYBR Green Assay, Migration

    42) Product Images from "Intermediates in V(D)J recombination: A stable RAG1/2 complex sequesters cleaved RSS ends"

    Article Title: Intermediates in V(D)J recombination: A stable RAG1/2 complex sequesters cleaved RSS ends

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    doi: 10.1073/pnas.221471198

    The effect of Mn 2+ on formation of the SEC. ( A ) Cleavage reactions of plasmid substrate were conducted in the presence of MgCl 2 (4 mM) with MnCl 2 (0.4 mM) being added either at the same time as MgCl 2 (0) or added 60 min after MgCl 2 (60), as indicated. The agarose gel stained with SYBR green I is shown. ( B and C ) Kinetic analyses were performed as described in Methods by using labeled 12 RSS plus unlabeled 23 RSS ( B ) or 12 RSS ( C ) at the concentrations indicated. Percent of labeled substrate converted into hairpin cleavage product at each time point was determined.
    Figure Legend Snippet: The effect of Mn 2+ on formation of the SEC. ( A ) Cleavage reactions of plasmid substrate were conducted in the presence of MgCl 2 (4 mM) with MnCl 2 (0.4 mM) being added either at the same time as MgCl 2 (0) or added 60 min after MgCl 2 (60), as indicated. The agarose gel stained with SYBR green I is shown. ( B and C ) Kinetic analyses were performed as described in Methods by using labeled 12 RSS plus unlabeled 23 RSS ( B ) or 12 RSS ( C ) at the concentrations indicated. Percent of labeled substrate converted into hairpin cleavage product at each time point was determined.

    Techniques Used: Size-exclusion Chromatography, Plasmid Preparation, Agarose Gel Electrophoresis, Staining, SYBR Green Assay, Labeling

    43) Product Images from "An mt+ gamete-specific nuclease that targets mt− chloroplasts during sexual reproduction in C. reinhardtii"

    Article Title: An mt+ gamete-specific nuclease that targets mt− chloroplasts during sexual reproduction in C. reinhardtii

    Journal: Genes & Development

    doi: 10.1101/gad.979902

    ( A ) Phase-contrast ( a ) and fluorescent ( b ) images of intact chloroplasts isolated from zygotes using an airbrush. To distinguish mt + and mt − chloroplasts, mocA84 mt + and 137c mt − gametes were crossed to form zygotes. Fluorescent images of SYBR Green I-stained gametes ( mocA84 mt + [ c ]; 137c mt − [ d ]) and a zygote ( mocA84 mt + × 137c mt − [ e ]). moc -type, wild-type cp nucleoids, and mt nucleoids are indicated by large arrows, small arrows, and white arrowheads, respectively. Phase contrast ( f,h,j,l,n ) and fluorescent images ( g,i,k,m,o ) of isolated chloroplasts stained by SYBR Green I. Intact mt + ( f,g,j,k ) and mt − ( h,i,l,m ) chloroplasts before ( f – i ) and after ( j – m ) active digestion, and fused chloroplasts ( n,o ) were observed. (Py) Pyrenoid. ( B ) Ca 2+ -dependent nuclease activity in chloroplasts detected by native-PAGE/in gelo assay. Chloroplasts were isolated from zygotes ( 137c mt + × 137c mt − ) at 30, 45, 60, and 90 min after mating. As a control, CBB-stained RuBisCO bands are shown under each lane. ( C ) Intact chloroplasts and frozen chloroplasts were treated with 0, 0.1, 1, and 10 mg/mL thermolysin. Their Ca 2+ nuclease activities were investigated by native-PAGE/in gelo assay.
    Figure Legend Snippet: ( A ) Phase-contrast ( a ) and fluorescent ( b ) images of intact chloroplasts isolated from zygotes using an airbrush. To distinguish mt + and mt − chloroplasts, mocA84 mt + and 137c mt − gametes were crossed to form zygotes. Fluorescent images of SYBR Green I-stained gametes ( mocA84 mt + [ c ]; 137c mt − [ d ]) and a zygote ( mocA84 mt + × 137c mt − [ e ]). moc -type, wild-type cp nucleoids, and mt nucleoids are indicated by large arrows, small arrows, and white arrowheads, respectively. Phase contrast ( f,h,j,l,n ) and fluorescent images ( g,i,k,m,o ) of isolated chloroplasts stained by SYBR Green I. Intact mt + ( f,g,j,k ) and mt − ( h,i,l,m ) chloroplasts before ( f – i ) and after ( j – m ) active digestion, and fused chloroplasts ( n,o ) were observed. (Py) Pyrenoid. ( B ) Ca 2+ -dependent nuclease activity in chloroplasts detected by native-PAGE/in gelo assay. Chloroplasts were isolated from zygotes ( 137c mt + × 137c mt − ) at 30, 45, 60, and 90 min after mating. As a control, CBB-stained RuBisCO bands are shown under each lane. ( C ) Intact chloroplasts and frozen chloroplasts were treated with 0, 0.1, 1, and 10 mg/mL thermolysin. Their Ca 2+ nuclease activities were investigated by native-PAGE/in gelo assay.

    Techniques Used: Isolation, SYBR Green Assay, Staining, Activity Assay, Clear Native PAGE

    ( A ) Phase-contrast ( a,c,e,g,i,k,m,o,q,r ) and fluorescent ( b,d,f,h,j,l,n,p,s,t ) images of SYBR Green I-stained gametes ( mocA84 mt + [ a,b,e,f ]; 137c mt − [ c,d,g,h ]), zygotes (30 min [ i – l ]; 90 min [ m – p ] after mating), and mt + ( q,s ) and mt − ( r,t ) chloroplasts isolated from zygotes at 30 min after mating. Gametes that are incubated for 10 d accumulate large quantities of starch granules in their chloroplasts (st+; a,b,g,h ), whereas gametes that are incubated for 2 d accumulate few starch granules (st−; c,d,e,f ). By crossing the two kinds of gametes, zygotes can be formed that accumulate large numbers of starch granules only in mt + chloroplasts ( i,j,m,n ) or mt − chloroplasts ( k,l,o,p ). The presence of a large amount of starch did not affect the active digestion of mt − cpDNA ( m – p ). Chloroplasts containing large amounts of starch granules were isolated and selectively separated by Percoll step-gradient centrifugation. The purity of separated chloroplasts was verified by fluorescence microscopy ( q – t ). (St) Starch granules. ( B ) Comparison of Ca 2+ -dependent nuclease activity in mt + and mt − chloroplasts using the native-PAGE/in gelo assay. The mt + and mt − chloroplasts that were isolated from zygotes at 30, 60, and 90 min after mating were analyzed. Experiments were repeated twice to ensure reproducibility. As a control, CBB-stained RuBisCO bands are shown under each lane.
    Figure Legend Snippet: ( A ) Phase-contrast ( a,c,e,g,i,k,m,o,q,r ) and fluorescent ( b,d,f,h,j,l,n,p,s,t ) images of SYBR Green I-stained gametes ( mocA84 mt + [ a,b,e,f ]; 137c mt − [ c,d,g,h ]), zygotes (30 min [ i – l ]; 90 min [ m – p ] after mating), and mt + ( q,s ) and mt − ( r,t ) chloroplasts isolated from zygotes at 30 min after mating. Gametes that are incubated for 10 d accumulate large quantities of starch granules in their chloroplasts (st+; a,b,g,h ), whereas gametes that are incubated for 2 d accumulate few starch granules (st−; c,d,e,f ). By crossing the two kinds of gametes, zygotes can be formed that accumulate large numbers of starch granules only in mt + chloroplasts ( i,j,m,n ) or mt − chloroplasts ( k,l,o,p ). The presence of a large amount of starch did not affect the active digestion of mt − cpDNA ( m – p ). Chloroplasts containing large amounts of starch granules were isolated and selectively separated by Percoll step-gradient centrifugation. The purity of separated chloroplasts was verified by fluorescence microscopy ( q – t ). (St) Starch granules. ( B ) Comparison of Ca 2+ -dependent nuclease activity in mt + and mt − chloroplasts using the native-PAGE/in gelo assay. The mt + and mt − chloroplasts that were isolated from zygotes at 30, 60, and 90 min after mating were analyzed. Experiments were repeated twice to ensure reproducibility. As a control, CBB-stained RuBisCO bands are shown under each lane.

    Techniques Used: SYBR Green Assay, Staining, Isolation, Incubation, Gradient Centrifugation, Fluorescence, Microscopy, Activity Assay, Clear Native PAGE

    ( A ) Phase-contrast ( a,c,e,g,i,k,m,o ) and fluorescent ( b,d,f,h,j,l,n,p ) images of living gametes ( 137c mt + [ e,f ]; 137c mt − [ c,d ]; mocA84 mt + [ a,b ]; mocA84 mt − [ g,h ]), and zygotes at 30 min ( i – l ) and 90 min ( m – p ) after mating for mocA84 mt + × 137c mt − ( i,j,m,n ), and 137c mt + × A84 mt − ( k,l,o,p ) crosses. The cells are stained with SYBR Green I. The yellow-green spheres (N) are cell nuclei. Chlorophyll autofluorescence is red. The yellow spots of the moc -type cp nucleoids, wild-type cp nucleoids, and the mitochondrial nucleoids are indicated by large arrows, small arrows, and white arrowheads, respectively. ( B ) Frequency distribution of the fluorescence of DAPI-stained moc -type nucleoids, quantified by VIMPCS at 30, 60, and 90 min after mating, and expressed in terms of cpDNA copy number. The fluorescence of mt + (white) and mt − (black) cp nucleoids in the mocA84 mt + × 137c mt − , and 137c mt + × A84 mt − crosses are shown.
    Figure Legend Snippet: ( A ) Phase-contrast ( a,c,e,g,i,k,m,o ) and fluorescent ( b,d,f,h,j,l,n,p ) images of living gametes ( 137c mt + [ e,f ]; 137c mt − [ c,d ]; mocA84 mt + [ a,b ]; mocA84 mt − [ g,h ]), and zygotes at 30 min ( i – l ) and 90 min ( m – p ) after mating for mocA84 mt + × 137c mt − ( i,j,m,n ), and 137c mt + × A84 mt − ( k,l,o,p ) crosses. The cells are stained with SYBR Green I. The yellow-green spheres (N) are cell nuclei. Chlorophyll autofluorescence is red. The yellow spots of the moc -type cp nucleoids, wild-type cp nucleoids, and the mitochondrial nucleoids are indicated by large arrows, small arrows, and white arrowheads, respectively. ( B ) Frequency distribution of the fluorescence of DAPI-stained moc -type nucleoids, quantified by VIMPCS at 30, 60, and 90 min after mating, and expressed in terms of cpDNA copy number. The fluorescence of mt + (white) and mt − (black) cp nucleoids in the mocA84 mt + × 137c mt − , and 137c mt + × A84 mt − crosses are shown.

    Techniques Used: Staining, SYBR Green Assay, Fluorescence

    44) Product Images from "Poly-γ-Glutamic Acid: Biodegradable Polymer for Potential Protection of Beneficial Viruses"

    Article Title: Poly-γ-Glutamic Acid: Biodegradable Polymer for Potential Protection of Beneficial Viruses

    Journal: Materials

    doi: 10.3390/ma9010028

    SYBR green I (2.5% v/v) stained samples were investigated under Fluorescence Microscopy: ( a ) T2 in PBS (non-formulated phage) well dispersed as represented by well-spaced bright particles as shown by red arrows; ( b ) T2 formulated with γ-PGA, clearly virion particles are associated with γ-PGA as shown within red ring. Scale bar, 10 μm.
    Figure Legend Snippet: SYBR green I (2.5% v/v) stained samples were investigated under Fluorescence Microscopy: ( a ) T2 in PBS (non-formulated phage) well dispersed as represented by well-spaced bright particles as shown by red arrows; ( b ) T2 formulated with γ-PGA, clearly virion particles are associated with γ-PGA as shown within red ring. Scale bar, 10 μm.

    Techniques Used: SYBR Green Assay, Staining, Fluorescence, Microscopy

    45) Product Images from "Development of Mitochondrial Loop-Mediated Isothermal Amplification for Detection of the Small Liver Fluke Opisthorchis viverrini (Opisthorchiidae; Trematoda; Platyhelminthes)"

    Article Title: Development of Mitochondrial Loop-Mediated Isothermal Amplification for Detection of the Small Liver Fluke Opisthorchis viverrini (Opisthorchiidae; Trematoda; Platyhelminthes)

    Journal: Journal of Clinical Microbiology

    doi: 10.1128/JCM.06277-11

    Sensitivity assessment of the LAMP assay for O. viverrini using serial dilutions of genomic DNA template and comparison with PCR (performed using outer primers F3 and B3). (A and B) Sensitivity analysis of the LAMP assay by visualization on 1.5% agarose stained with ethidium bromide (A) or under UV light after the addition of SYBR green I (B). (C) Sensitivity analysis of the conventional PCR revealed on 1% agarose (arrow indicates a 216-bp DNA fragment at the highest template dilution for PCR performance). Lanes: M, 1-kb ladder marker (numbers on the left are sizes in base pairs); (-), no-DNA template (water added) as a negative control; (+), 100 ng template (positive control), 10, 10 ng; 1, 1 ng; −1: 100 pg; −2, 10 pg; −3, 1 pg; −4, 100 fg; −5, 10 fg.
    Figure Legend Snippet: Sensitivity assessment of the LAMP assay for O. viverrini using serial dilutions of genomic DNA template and comparison with PCR (performed using outer primers F3 and B3). (A and B) Sensitivity analysis of the LAMP assay by visualization on 1.5% agarose stained with ethidium bromide (A) or under UV light after the addition of SYBR green I (B). (C) Sensitivity analysis of the conventional PCR revealed on 1% agarose (arrow indicates a 216-bp DNA fragment at the highest template dilution for PCR performance). Lanes: M, 1-kb ladder marker (numbers on the left are sizes in base pairs); (-), no-DNA template (water added) as a negative control; (+), 100 ng template (positive control), 10, 10 ng; 1, 1 ng; −1: 100 pg; −2, 10 pg; −3, 1 pg; −4, 100 fg; −5, 10 fg.

    Techniques Used: Lamp Assay, Polymerase Chain Reaction, Staining, SYBR Green Assay, Marker, Negative Control, Positive Control

    Results of specific LAMP assays for detection of O. viverrini using DNA material from mixed metacercariae extracted from freshwater fish and eggs from feces of humans. (A) Visualization on 1.5% agarose stained with ethidium bromide; (B) with SYBR green I under UV light. Lanes: M, 1-kb ladder marker (numbers on the left are sizes in base pairs); (-), LAMP negative control (no DNA template); 1 through 3, DNA of possibly mixed metacercariae from muscles of fish caught in ponds where O. viverrini is endemic; 4 through 7, LAMP products from DNA of eggs, possibly of more than one species, extracted from individual stool samples from humans (after raw fish consumption) in a village where O. viverrini is endemic.
    Figure Legend Snippet: Results of specific LAMP assays for detection of O. viverrini using DNA material from mixed metacercariae extracted from freshwater fish and eggs from feces of humans. (A) Visualization on 1.5% agarose stained with ethidium bromide; (B) with SYBR green I under UV light. Lanes: M, 1-kb ladder marker (numbers on the left are sizes in base pairs); (-), LAMP negative control (no DNA template); 1 through 3, DNA of possibly mixed metacercariae from muscles of fish caught in ponds where O. viverrini is endemic; 4 through 7, LAMP products from DNA of eggs, possibly of more than one species, extracted from individual stool samples from humans (after raw fish consumption) in a village where O. viverrini is endemic.

    Techniques Used: Fluorescence In Situ Hybridization, Staining, SYBR Green Assay, Marker, Negative Control

    46) Product Images from "Loop-mediated isothermal amplification of DNA"

    Article Title: Loop-mediated isothermal amplification of DNA

    Journal: Nucleic Acids Research

    doi:

    Sensitivity of LAMP. ( A ) Time course of the LAMP reaction with various amounts of HBV DNA. Various numbers of copies of HBV DNA were amplified by LAMP. At various times, the reaction was terminated and the amounts of products quantified by measuring fluorescence intensity of SYBR Green I. ( B ) Requirements for primers in the LAMP reaction. Sixty copies of HBV DNA were amplified by LAMP with omission of one or two of the primers. The products were electrophoresed in 2% agarose gels and stained. –, the corresponding primer was omitted from the reaction; B2 and F2, BIP and FIP were replaced by B2 and F2, respectively, which do not contain B1c and F1c and, therefore, are unable to form the looped out structure. ( C ) The effect of the presence of genomic DNA on sensitivity. Various numbers of copies of HBV DNA were amplified at 60°C for 60 min in the absence or presence of 100 ng of human genomic DNA and the products separated by gel electrophoresis. Lane M, 100 bp ladder size markers (TaKaRa); lanes 1–4, LAMP carried out in the absence of human genomic DNA; lanes 5–8, LAMP carried out in the presence of 100 ng genomic DNA; lanes 1 and 5, LAMP without HBV DNA; lanes 2 and 6, with six copies; lanes 3 and 7, with 60 copies; lanes 4 and 8, with 600 copies of HBV DNA. Lanes 9 and 10 are Ear I digests (1/5 vol) of the same amplified DNAs as in lanes 2 and 6, respectively. ( D ) Nested PCR of HBV DNA under similar conditions. Single and nested PCR reactions were performed in a 50 µl reaction mixture containing 2.5 U AmpliTaq Gold (PE Biosystems), 0.2 µM each primer (first PCR, HBVB2/HBVF2 or HBVB1/HBVF1; second PCR, HBVB1/HBVF1), 1× GeneAmp PCR buffer (10 mM Tris–HCl, pH 8.3, 50 mM KCl, 1.5 mM MgCl 2 , 0.01% gelatin) and 0.2 mM dNTPs. The sequences of the primers used were 5′-CCAACCTCTTGTCCTCCAA-3′ for HBVB2, 5′-GACAAACGGGCAACATACCTT-3′ for HBVF2, 5′-GGATGTGTCTGCGGCGTTTTATC-3′ for HBVB1 and 5′-AGAAGATGAGGCATAGCAGCAGG-3′ for HBVF1. Both the first and second round nested PCRs were carried out as follows: preincubation at 95°C for 10 min; 40 cycles each of 30 s at 95°C, 30 s at 60°C and 1 min at 72°C. One microliter of the first PCR products (HBVB2/HBVF2) was subjected to second PCR. An aliquot of 10 µl of the reaction products was analyzed by 4% agarose gel (0.5× TBE) electrophoresis followed by staining with SYBR Green I.
    Figure Legend Snippet: Sensitivity of LAMP. ( A ) Time course of the LAMP reaction with various amounts of HBV DNA. Various numbers of copies of HBV DNA were amplified by LAMP. At various times, the reaction was terminated and the amounts of products quantified by measuring fluorescence intensity of SYBR Green I. ( B ) Requirements for primers in the LAMP reaction. Sixty copies of HBV DNA were amplified by LAMP with omission of one or two of the primers. The products were electrophoresed in 2% agarose gels and stained. –, the corresponding primer was omitted from the reaction; B2 and F2, BIP and FIP were replaced by B2 and F2, respectively, which do not contain B1c and F1c and, therefore, are unable to form the looped out structure. ( C ) The effect of the presence of genomic DNA on sensitivity. Various numbers of copies of HBV DNA were amplified at 60°C for 60 min in the absence or presence of 100 ng of human genomic DNA and the products separated by gel electrophoresis. Lane M, 100 bp ladder size markers (TaKaRa); lanes 1–4, LAMP carried out in the absence of human genomic DNA; lanes 5–8, LAMP carried out in the presence of 100 ng genomic DNA; lanes 1 and 5, LAMP without HBV DNA; lanes 2 and 6, with six copies; lanes 3 and 7, with 60 copies; lanes 4 and 8, with 600 copies of HBV DNA. Lanes 9 and 10 are Ear I digests (1/5 vol) of the same amplified DNAs as in lanes 2 and 6, respectively. ( D ) Nested PCR of HBV DNA under similar conditions. Single and nested PCR reactions were performed in a 50 µl reaction mixture containing 2.5 U AmpliTaq Gold (PE Biosystems), 0.2 µM each primer (first PCR, HBVB2/HBVF2 or HBVB1/HBVF1; second PCR, HBVB1/HBVF1), 1× GeneAmp PCR buffer (10 mM Tris–HCl, pH 8.3, 50 mM KCl, 1.5 mM MgCl 2 , 0.01% gelatin) and 0.2 mM dNTPs. The sequences of the primers used were 5′-CCAACCTCTTGTCCTCCAA-3′ for HBVB2, 5′-GACAAACGGGCAACATACCTT-3′ for HBVF2, 5′-GGATGTGTCTGCGGCGTTTTATC-3′ for HBVB1 and 5′-AGAAGATGAGGCATAGCAGCAGG-3′ for HBVF1. Both the first and second round nested PCRs were carried out as follows: preincubation at 95°C for 10 min; 40 cycles each of 30 s at 95°C, 30 s at 60°C and 1 min at 72°C. One microliter of the first PCR products (HBVB2/HBVF2) was subjected to second PCR. An aliquot of 10 µl of the reaction products was analyzed by 4% agarose gel (0.5× TBE) electrophoresis followed by staining with SYBR Green I.

    Techniques Used: Amplification, Fluorescence, SYBR Green Assay, Staining, Nucleic Acid Electrophoresis, Nested PCR, Polymerase Chain Reaction, Agarose Gel Electrophoresis, Electrophoresis

    Detection of PSA mRNA by reverse transcription-coupled LAMP (RT-LAMP). Various numbers of LNCaP cells were mixed with 10 6 PSA-non-producing K562 cells and total RNA was extracted. RT-LAMP was carried out in the same reaction mixture as for M13mp18 DNA amplification except that 1.6 µM each PSAFIP and PSABIP, 0.2 µM each PSAF3 and PSAB3, 0.8 M betaine, 5 mM DTT, 16 U Bst polymerase, 100 U ReverTra Ace (Toyobo) and 5 µg of extracted RNA were used. All the above components were mixed at once on ice and were incubated at 65°C for 45 min. The products were electrophoresed in 2% agarose gel followed by SYBR Green I staining. + and –, RT-LAMP carried out in the presence and absence of Bst DNA polymerase or ReverTra Ace, respectively. Lanes 8 and 9, the same products (1/5 vol) as in lanes 6 and 7, respectively, but digested with Sau 3AI; lane M, 100 bp ladder (New England Biolabs).
    Figure Legend Snippet: Detection of PSA mRNA by reverse transcription-coupled LAMP (RT-LAMP). Various numbers of LNCaP cells were mixed with 10 6 PSA-non-producing K562 cells and total RNA was extracted. RT-LAMP was carried out in the same reaction mixture as for M13mp18 DNA amplification except that 1.6 µM each PSAFIP and PSABIP, 0.2 µM each PSAF3 and PSAB3, 0.8 M betaine, 5 mM DTT, 16 U Bst polymerase, 100 U ReverTra Ace (Toyobo) and 5 µg of extracted RNA were used. All the above components were mixed at once on ice and were incubated at 65°C for 45 min. The products were electrophoresed in 2% agarose gel followed by SYBR Green I staining. + and –, RT-LAMP carried out in the presence and absence of Bst DNA polymerase or ReverTra Ace, respectively. Lanes 8 and 9, the same products (1/5 vol) as in lanes 6 and 7, respectively, but digested with Sau 3AI; lane M, 100 bp ladder (New England Biolabs).

    Techniques Used: Amplification, Incubation, Agarose Gel Electrophoresis, SYBR Green Assay, Staining

    Restriction analysis and Southern blot hybridization of the amplified M13mp18 DNA. ( A ) Electrophoretic analysis of the LAMP amplified M13mp18 product. Six hundred copies of M13mp18 DNA were amplified by LAMP with the specific primers designed on the sequences shown in Figure 2 and run on a 2% agarose gel followed by SYBR Green I staining. Lane M, 100 bp ladder used as size marker (New England Biolabs); lane 1, M13mpl8 DNA digested with Pvu II; lane 2, LAMP without Bst DNA polymerase; lane 3, LAMP without target M13 DNA; lane 4, complete LAMP; lanes 5–7, complete LAMP products after digestion with Bam HI, Pst I and Pvu II, respectively (one fifth of the digests were loaded). (B–D) Southern blot analysis of the LAMP products. The 2% agarose gel shown in (A) was used for Southern blot hybridization with M13-281 DNA ( B ), M13-333 DNA ( C ) and M13BIP ( D ) as probes. ( E ) Alkaline agarose gel electrophoresis of the LAMP products. Lane m, λ DNA Hin dIII digests; lane 4, the same sample as in (A).
    Figure Legend Snippet: Restriction analysis and Southern blot hybridization of the amplified M13mp18 DNA. ( A ) Electrophoretic analysis of the LAMP amplified M13mp18 product. Six hundred copies of M13mp18 DNA were amplified by LAMP with the specific primers designed on the sequences shown in Figure 2 and run on a 2% agarose gel followed by SYBR Green I staining. Lane M, 100 bp ladder used as size marker (New England Biolabs); lane 1, M13mpl8 DNA digested with Pvu II; lane 2, LAMP without Bst DNA polymerase; lane 3, LAMP without target M13 DNA; lane 4, complete LAMP; lanes 5–7, complete LAMP products after digestion with Bam HI, Pst I and Pvu II, respectively (one fifth of the digests were loaded). (B–D) Southern blot analysis of the LAMP products. The 2% agarose gel shown in (A) was used for Southern blot hybridization with M13-281 DNA ( B ), M13-333 DNA ( C ) and M13BIP ( D ) as probes. ( E ) Alkaline agarose gel electrophoresis of the LAMP products. Lane m, λ DNA Hin dIII digests; lane 4, the same sample as in (A).

    Techniques Used: Southern Blot, Hybridization, Amplification, Agarose Gel Electrophoresis, SYBR Green Assay, Staining, Marker

    47) Product Images from "Rapid and sensitive diagnoses of dry root rot pathogen of chickpea (Rhizoctonia bataticola (Taub.) Butler) using loop-mediated isothermal amplification assay"

    Article Title: Rapid and sensitive diagnoses of dry root rot pathogen of chickpea (Rhizoctonia bataticola (Taub.) Butler) using loop-mediated isothermal amplification assay

    Journal: Scientific Reports

    doi: 10.1038/srep42737

    Sensitivity of LAMP assay vs. conventional PCR for detection of Rhizoctonia bataticola using similar concentration of DNA template. ( a ) Visual assessment of LAMP assay using SYBR Green I. ( b ) LAMP assay on the basis of 2% agarose gel electrophoresis. ( c ) Result of conventional PCR using ITS1 and ITS4 primers. M GeneRuler TM 100 bp Plus DNA Ladder; lane 10 1 –10 −7 indicated the DNA concentration in LAMP reaction starting from 10 ng (10 1 ng) to subsequent 10 fold diluted DNA up to 0.1 fg (10 −7 ng), respectively.
    Figure Legend Snippet: Sensitivity of LAMP assay vs. conventional PCR for detection of Rhizoctonia bataticola using similar concentration of DNA template. ( a ) Visual assessment of LAMP assay using SYBR Green I. ( b ) LAMP assay on the basis of 2% agarose gel electrophoresis. ( c ) Result of conventional PCR using ITS1 and ITS4 primers. M GeneRuler TM 100 bp Plus DNA Ladder; lane 10 1 –10 −7 indicated the DNA concentration in LAMP reaction starting from 10 ng (10 1 ng) to subsequent 10 fold diluted DNA up to 0.1 fg (10 −7 ng), respectively.

    Techniques Used: Lamp Assay, Polymerase Chain Reaction, Concentration Assay, SYBR Green Assay, Agarose Gel Electrophoresis

    48) Product Images from "Rapid detection of Opisthorchis viverrini copro-DNA using loop-mediated isothermal amplification (LAMP)"

    Article Title: Rapid detection of Opisthorchis viverrini copro-DNA using loop-mediated isothermal amplification (LAMP)

    Journal: Parasitology international

    doi: 10.1016/j.parint.2011.08.009

    Real-time monitoring of LAMP reaction. The generated fluorescence intensity of DNA binding SYBR Green I was monitored at every 1 minute. Template DNAs were prepared from 10 1 to 10 -5 ng/μL. Distilled water (D.W.) was used as a negative control.
    Figure Legend Snippet: Real-time monitoring of LAMP reaction. The generated fluorescence intensity of DNA binding SYBR Green I was monitored at every 1 minute. Template DNAs were prepared from 10 1 to 10 -5 ng/μL. Distilled water (D.W.) was used as a negative control.

    Techniques Used: Generated, Fluorescence, Binding Assay, SYBR Green Assay, Negative Control

    49) Product Images from "Antimicrobial Activity of Bee Venom and Melittin against Borrelia burgdorferi"

    Article Title: Antimicrobial Activity of Bee Venom and Melittin against Borrelia burgdorferi

    Journal: Antibiotics

    doi: 10.3390/antibiotics6040031

    Representative Live/Dead staining images of B. burgdorferi 7-day recovery cultures following treatment with different antimicrobial agents. Cells were stained with SYBR Green I/PI as outlined in the Material and Methods and representative images were taken at 100× magnification. Panel ( A ) Borrelia culture treated only with PBS was used as a negative control. Panel ( B ) Doxycycline (DOXY) treated, Panel ( C ) Cefoperazone (CEFO) treated, Panel ( D ) Daptomycin (DAPTO) treated and Panel ( E ) Three-antibiotic combination (D + C + D). Panels ( F – H ) Bee venom (BV) was used in increasing concentrations while Panels ( I – K ) depict melittin (M) treated cells at different concentrations. Live cells are stained with green color while dead cells are stained red. Scale bar: 100 μm.
    Figure Legend Snippet: Representative Live/Dead staining images of B. burgdorferi 7-day recovery cultures following treatment with different antimicrobial agents. Cells were stained with SYBR Green I/PI as outlined in the Material and Methods and representative images were taken at 100× magnification. Panel ( A ) Borrelia culture treated only with PBS was used as a negative control. Panel ( B ) Doxycycline (DOXY) treated, Panel ( C ) Cefoperazone (CEFO) treated, Panel ( D ) Daptomycin (DAPTO) treated and Panel ( E ) Three-antibiotic combination (D + C + D). Panels ( F – H ) Bee venom (BV) was used in increasing concentrations while Panels ( I – K ) depict melittin (M) treated cells at different concentrations. Live cells are stained with green color while dead cells are stained red. Scale bar: 100 μm.

    Techniques Used: Staining, SYBR Green Assay, Negative Control

    The effects of various antimicrobial agents on B. burgdorferi as determined by SYBR Green I/PI assay Panel ( A ) or direct counting assay Panel ( B ). Doxycycline, Cefoperazone, Daptomycin and their combination (D + C + D) as well as different concentrations of bee venom and melittin were tested on B. burgdorferi logarithmic phase (spirochetes) culture and stationary phase (persisters) cultures as well as in 7-day recovery subculture as described previously [ 6 , 7 , 8 ]. Significance against sterile PBS buffer (control vehicle) with the p value of
    Figure Legend Snippet: The effects of various antimicrobial agents on B. burgdorferi as determined by SYBR Green I/PI assay Panel ( A ) or direct counting assay Panel ( B ). Doxycycline, Cefoperazone, Daptomycin and their combination (D + C + D) as well as different concentrations of bee venom and melittin were tested on B. burgdorferi logarithmic phase (spirochetes) culture and stationary phase (persisters) cultures as well as in 7-day recovery subculture as described previously [ 6 , 7 , 8 ]. Significance against sterile PBS buffer (control vehicle) with the p value of

    Techniques Used: SYBR Green Assay

    Representative Live/Dead staining images of B. burgdorferi stationary phase persister cultures following treatment with different antimicrobial agents. Cells were stained with SYBR Green I/PI as outlined in the Methods and representative images were taken at 100× magnification. Panel ( A ) Borrelia culture treated only with PBS was used as a negative control. Panel ( B ) Doxycycline (DOXY) treated, Panel ( C ) Cefoperazone (CEFO) treated, Panel ( D ) Daptomycin (DAPTO) treated and Panel ( E ) Three-antibiotic combination (D + C + D). Panels ( F – H ) Bee venom (BV) was used in increasing concentrations while Panels ( I – K ) depicts melittin (M) treated cells. Live cells are stained with green color while dead cells are stained red. Scale bar: 100 μm.
    Figure Legend Snippet: Representative Live/Dead staining images of B. burgdorferi stationary phase persister cultures following treatment with different antimicrobial agents. Cells were stained with SYBR Green I/PI as outlined in the Methods and representative images were taken at 100× magnification. Panel ( A ) Borrelia culture treated only with PBS was used as a negative control. Panel ( B ) Doxycycline (DOXY) treated, Panel ( C ) Cefoperazone (CEFO) treated, Panel ( D ) Daptomycin (DAPTO) treated and Panel ( E ) Three-antibiotic combination (D + C + D). Panels ( F – H ) Bee venom (BV) was used in increasing concentrations while Panels ( I – K ) depicts melittin (M) treated cells. Live cells are stained with green color while dead cells are stained red. Scale bar: 100 μm.

    Techniques Used: Staining, SYBR Green Assay, Negative Control

    Representative Live/Dead staining images of B. burgdorferi log phase spirochetal cultures treated with different antimicrobial agents. Cells were stained with SYBR Green I/PI as outlined in the Methods and representative images were taken at 100× magnification. Panel ( A ) Borrelia culture treated only with PBS was used as a negative control. Panel ( B ) Doxycycline (DOXY) treated; Panel ( C ) Cefoperazone (CEFO) treated; Panel ( D ) Daptomycin (DAPTO) treated and Panel ( E ) Three-antibiotic combination (D + C + D). Panels ( F – H ) Bee venom (BV) was used in increasing concentrations while Panels ( I – K ) depicts melittin (M) treated cells. Live cells are stained with green color while dead cells are stained red. Scale bar: 100 μm.
    Figure Legend Snippet: Representative Live/Dead staining images of B. burgdorferi log phase spirochetal cultures treated with different antimicrobial agents. Cells were stained with SYBR Green I/PI as outlined in the Methods and representative images were taken at 100× magnification. Panel ( A ) Borrelia culture treated only with PBS was used as a negative control. Panel ( B ) Doxycycline (DOXY) treated; Panel ( C ) Cefoperazone (CEFO) treated; Panel ( D ) Daptomycin (DAPTO) treated and Panel ( E ) Three-antibiotic combination (D + C + D). Panels ( F – H ) Bee venom (BV) was used in increasing concentrations while Panels ( I – K ) depicts melittin (M) treated cells. Live cells are stained with green color while dead cells are stained red. Scale bar: 100 μm.

    Techniques Used: Staining, SYBR Green Assay, Negative Control

    Representative Live/Dead images of the viability of attached Borrelia biofilms following treatment with different antimicrobial agents. Biofilms were stained with SYBR Green I and PI as outlined in the Material and Methods and representative images were taken at 100× magnification. Panel ( A ) Borrelia culture treated only with PBS was used as a negative control. Panel ( B ) Doxycycline (DOXY) treated, Panel ( C ) Cefoperazone (CEFO) treated, Panel ( D ) Daptomycin (DAPTO) treated and Panel ( E ) Three-antibiotic combination (D + C + D). Panels ( F – H ) Bee venom (BV) was used in increasing concentrations while Panels ( I – K ) depict melittin treated cells at different concentration. Live cells are stained with green color while dead cells are stained red. Scale bar: 100 μm.
    Figure Legend Snippet: Representative Live/Dead images of the viability of attached Borrelia biofilms following treatment with different antimicrobial agents. Biofilms were stained with SYBR Green I and PI as outlined in the Material and Methods and representative images were taken at 100× magnification. Panel ( A ) Borrelia culture treated only with PBS was used as a negative control. Panel ( B ) Doxycycline (DOXY) treated, Panel ( C ) Cefoperazone (CEFO) treated, Panel ( D ) Daptomycin (DAPTO) treated and Panel ( E ) Three-antibiotic combination (D + C + D). Panels ( F – H ) Bee venom (BV) was used in increasing concentrations while Panels ( I – K ) depict melittin treated cells at different concentration. Live cells are stained with green color while dead cells are stained red. Scale bar: 100 μm.

    Techniques Used: Staining, SYBR Green Assay, Negative Control, Concentration Assay

    50) Product Images from "The YlmG protein has a conserved function related to the distribution of nucleoids in chloroplasts and cyanobacteria"

    Article Title: The YlmG protein has a conserved function related to the distribution of nucleoids in chloroplasts and cyanobacteria

    Journal: BMC Plant Biology

    doi: 10.1186/1471-2229-10-57

    Effects of the overexpression and knockdown of AtYLMG1-1 on the morphology of the chloroplast nucleoids . (A) Morphology of the chloroplast nucleoids in the overexpresser and the knockdown lines. Expanding leaf or the basal part of expanding leaf cells of the wild type (WT), the AtYLMG1-1 knockdown line (AS), and the AtYLMG1-1 overexpresser (OX) were stained with DAPI. The white portion indicates DAPI fluorescence showing the localization of DNA. Nuclei (N) are also observed in some panels. Magnified images are also shown in the lower panels. Bars = 5 μm. All images were obtained with the same exposure time. (B) Morphology of the nucleoids in dividing chloroplasts. Young emerging leaves of the wild type were stained with SYBR GREEN I. The white portion indicates the SYBR GREEN I fluorescence showing the localization of DNA. Arrowheads indicate dividing chloroplasts. Other dividing chloroplasts are also shown in the right panels. Bars = 10 μm. (C) Comparison of the quantity of chloroplast DNA by DNA-blot analysis. Total genome DNA of the wild type (WT), the AtYLMG1-1 overexpresser (OX), and the AtYLMG1-1 knockdown line (AS) was extracted and then was digested with Hin dIII. Three micrograms of digested DNA were loaded in each lane. Chloroplast DNA (cp) was detected with a psbA probe and nuclear DNA (nu) was detected with a PsbO probe. Nuclear DNA was detected as the quantitative control. (D) Morphology of the chloroplast nucleoids in ftsZ2-1 , arc5 , and arc6 mutants. Mature leaves of the ftsZ2-1 , arc5 , and arc6 mutants were stained with DAPI. The white portion indicates DAPI fluorescence showing the localization of DNA. Bars = 5 μm. All images were obtained with the same exposure time.
    Figure Legend Snippet: Effects of the overexpression and knockdown of AtYLMG1-1 on the morphology of the chloroplast nucleoids . (A) Morphology of the chloroplast nucleoids in the overexpresser and the knockdown lines. Expanding leaf or the basal part of expanding leaf cells of the wild type (WT), the AtYLMG1-1 knockdown line (AS), and the AtYLMG1-1 overexpresser (OX) were stained with DAPI. The white portion indicates DAPI fluorescence showing the localization of DNA. Nuclei (N) are also observed in some panels. Magnified images are also shown in the lower panels. Bars = 5 μm. All images were obtained with the same exposure time. (B) Morphology of the nucleoids in dividing chloroplasts. Young emerging leaves of the wild type were stained with SYBR GREEN I. The white portion indicates the SYBR GREEN I fluorescence showing the localization of DNA. Arrowheads indicate dividing chloroplasts. Other dividing chloroplasts are also shown in the right panels. Bars = 10 μm. (C) Comparison of the quantity of chloroplast DNA by DNA-blot analysis. Total genome DNA of the wild type (WT), the AtYLMG1-1 overexpresser (OX), and the AtYLMG1-1 knockdown line (AS) was extracted and then was digested with Hin dIII. Three micrograms of digested DNA were loaded in each lane. Chloroplast DNA (cp) was detected with a psbA probe and nuclear DNA (nu) was detected with a PsbO probe. Nuclear DNA was detected as the quantitative control. (D) Morphology of the chloroplast nucleoids in ftsZ2-1 , arc5 , and arc6 mutants. Mature leaves of the ftsZ2-1 , arc5 , and arc6 mutants were stained with DAPI. The white portion indicates DAPI fluorescence showing the localization of DNA. Bars = 5 μm. All images were obtained with the same exposure time.

    Techniques Used: Over Expression, Staining, Fluorescence, SYBR Green Assay

    51) Product Images from "Vertical and Seasonal Variations of Bacterioplankton Subgroups with Different Nucleic Acid Contents: Possible Regulation by Phosphorus †"

    Article Title: Vertical and Seasonal Variations of Bacterioplankton Subgroups with Different Nucleic Acid Contents: Possible Regulation by Phosphorus †

    Journal:

    doi: 10.1128/AEM.71.10.5828-5836.2005

    Example of a flow cytogram of bacterioplankton in Lake Biwa (the sample was collected on 15 October 2001 at a depth of 40 m at station 5). Side scatter (SSC) and fluorescence (FL1) emitted from SYBR Green I bound to bacterial nucleic acid differentiated
    Figure Legend Snippet: Example of a flow cytogram of bacterioplankton in Lake Biwa (the sample was collected on 15 October 2001 at a depth of 40 m at station 5). Side scatter (SSC) and fluorescence (FL1) emitted from SYBR Green I bound to bacterial nucleic acid differentiated

    Techniques Used: Flow Cytometry, Fluorescence, SYBR Green Assay

    52) Product Images from "Nonlinear electrophoretic response yields a unique parameter for separation of biomolecules"

    Article Title: Nonlinear electrophoretic response yields a unique parameter for separation of biomolecules

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    doi: 10.1073/pnas.0907402106

    SCODA concentration sequence. Time-lapse sequence showing concentration of SYBR Green I–stained pUC19 DNA (2.7 kb) from a homogeneous solution of 0.2 ng/μL of DNA in 1% agarose and 0.25× TBE, to a 750-μm-diameter spot. Images are taken at 10-min intervals, for a total run time of 60 min at a SCODA field of 250 V/cm (maximum field in SCODA gel). The concentration of DNA in the focused spot is estimated to be 100–200 ng/μL. ( A ) Diagram of dipole and quadrupole SCODA field lines. ( B–H ) SCODA duration, in minutes: B = 0, C = 10, D = 20, E = 30, F = 40, G = 50, and H = 60. Camera exposure is reduced to avoid saturation from increasing fluorescence intensity over the course of concentration. Exposure times, in milliseconds: B = 1,000, C = 750, D = 250, E = 50, F = 50, G = 10, and H = 10.
    Figure Legend Snippet: SCODA concentration sequence. Time-lapse sequence showing concentration of SYBR Green I–stained pUC19 DNA (2.7 kb) from a homogeneous solution of 0.2 ng/μL of DNA in 1% agarose and 0.25× TBE, to a 750-μm-diameter spot. Images are taken at 10-min intervals, for a total run time of 60 min at a SCODA field of 250 V/cm (maximum field in SCODA gel). The concentration of DNA in the focused spot is estimated to be 100–200 ng/μL. ( A ) Diagram of dipole and quadrupole SCODA field lines. ( B–H ) SCODA duration, in minutes: B = 0, C = 10, D = 20, E = 30, F = 40, G = 50, and H = 60. Camera exposure is reduced to avoid saturation from increasing fluorescence intensity over the course of concentration. Exposure times, in milliseconds: B = 1,000, C = 750, D = 250, E = 50, F = 50, G = 10, and H = 10.

    Techniques Used: Synchronous Coefficient of Drag Alteration, Concentration Assay, Sequencing, SYBR Green Assay, Staining, Fluorescence

    53) Product Images from "Rapid detection of contagious ecthyma by loop-mediated isothermal amplification and epidemiology in Jilin Province China"

    Article Title: Rapid detection of contagious ecthyma by loop-mediated isothermal amplification and epidemiology in Jilin Province China

    Journal: The Journal of Veterinary Medical Science

    doi: 10.1292/jvms.15-0340

    Establishment of the LAMP reaction for orf virus. A: SYBR Green I staining. 1: Negative control. 2: ORFV genomic DNA. 3: Standard plasmid. B: Agarose gel analysis. M: 100 bp ladder marker. N: Negative control. 1: ORFV genomic DNA. 2: Standard plasmid.
    Figure Legend Snippet: Establishment of the LAMP reaction for orf virus. A: SYBR Green I staining. 1: Negative control. 2: ORFV genomic DNA. 3: Standard plasmid. B: Agarose gel analysis. M: 100 bp ladder marker. N: Negative control. 1: ORFV genomic DNA. 2: Standard plasmid.

    Techniques Used: SYBR Green Assay, Staining, Negative Control, Plasmid Preparation, Agarose Gel Electrophoresis, Marker

    The sensitivity of LAMP and PCR. The panels show the DNA templates used at 0–10 8 copies/reaction of the ORFV standard plasmid. M:100 bp ladder marker. A, B: The sensitivity of LAMP with SYBR Green I staining and agarose gel analysis. C: The sensitivity of PCR with agarose gel analysis.
    Figure Legend Snippet: The sensitivity of LAMP and PCR. The panels show the DNA templates used at 0–10 8 copies/reaction of the ORFV standard plasmid. M:100 bp ladder marker. A, B: The sensitivity of LAMP with SYBR Green I staining and agarose gel analysis. C: The sensitivity of PCR with agarose gel analysis.

    Techniques Used: Polymerase Chain Reaction, Plasmid Preparation, Marker, SYBR Green Assay, Staining, Agarose Gel Electrophoresis

    54) Product Images from "Bacterial Communities of Diatoms Display Strong Conservation Across Strains and Time"

    Article Title: Bacterial Communities of Diatoms Display Strong Conservation Across Strains and Time

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2018.00659

    Growth of A3 and its microbiome. Cultures were acclimated as described in the Section “Materials and Methods.” Diatom growth (black circles) with its microbiome was monitored using relative fluorescence and bacterial growth with the diatom (red squares, solid line) or without the diatom (red squares, dashed line) was monitored using microscopy counts of Sybr Green I stained cells. Error bars represent standard deviation (SD) of six cultures.
    Figure Legend Snippet: Growth of A3 and its microbiome. Cultures were acclimated as described in the Section “Materials and Methods.” Diatom growth (black circles) with its microbiome was monitored using relative fluorescence and bacterial growth with the diatom (red squares, solid line) or without the diatom (red squares, dashed line) was monitored using microscopy counts of Sybr Green I stained cells. Error bars represent standard deviation (SD) of six cultures.

    Techniques Used: Fluorescence, Microscopy, SYBR Green Assay, Staining, Standard Deviation

    55) Product Images from "Direct loop-mediated isothermal amplification assay for on-site detection of Staphylococcus aureus"

    Article Title: Direct loop-mediated isothermal amplification assay for on-site detection of Staphylococcus aureus

    Journal: FEMS Microbiology Letters

    doi: 10.1093/femsle/fny092

    Sensitivity test of S. aureus by different LAMP assays. Visual LAMP detection of S. aureus via SYBR Green ® I staining. The LAMP assay was performed using DNA extracted by boiling the samples ( A ), and commercial kit ( B ). Lane 1–7, LAMP assays using DNA templates extracted from S. aureus cell of 7.6 × 10 6 CFU/mL, 7.6 × 10 5 CFU/mL, 7.6 × 10 4 CFU/mL, 7.6 × 10 3 CFU/mL, 7.6 × 10 2 CFU/mL, 7.6 × 10 1 CFU/mL, and 7.6 × 10 0 CFU/mL. Lane 8, negative control, in which no DNA template was added in the reaction mixture. Each assay was carried out in triplicate.
    Figure Legend Snippet: Sensitivity test of S. aureus by different LAMP assays. Visual LAMP detection of S. aureus via SYBR Green ® I staining. The LAMP assay was performed using DNA extracted by boiling the samples ( A ), and commercial kit ( B ). Lane 1–7, LAMP assays using DNA templates extracted from S. aureus cell of 7.6 × 10 6 CFU/mL, 7.6 × 10 5 CFU/mL, 7.6 × 10 4 CFU/mL, 7.6 × 10 3 CFU/mL, 7.6 × 10 2 CFU/mL, 7.6 × 10 1 CFU/mL, and 7.6 × 10 0 CFU/mL. Lane 8, negative control, in which no DNA template was added in the reaction mixture. Each assay was carried out in triplicate.

    Techniques Used: SYBR Green Assay, Staining, Lamp Assay, Negative Control

    56) Product Images from "Inducing controlled cell cycle arrest and re-entry during asexual proliferation of Plasmodium falciparum malaria parasites"

    Article Title: Inducing controlled cell cycle arrest and re-entry during asexual proliferation of Plasmodium falciparum malaria parasites

    Journal: Scientific Reports

    doi: 10.1038/s41598-018-34964-w

    DFMO arrest and putrescine reversal associated to G1/S cell cycle control in intraerythrocytic P. falciparum parasites. In all instances, ■ refers to control parasites, ▼ refers to DFMO-treated parasites, and ● refers to putrescine-reversed parasites. ( a ) DFMO dose-response curves of asexual P . falciparum 3D7 proliferation over 96 h (initiated with ring-stage parasites, 1% haematocrit, 1% parasitaemia) at 37 °C in the absence or presence of 2 mM putrescine. Proliferation is expressed relative to untreated controls, with data averaged from n = 6 biological replicates and shown ± S.E. ( b ) Synchronised P . falciparum 3D7 cultures were treated with DFMO alone (IC 90 ) or with putrescine (2 mM, after 24 h DFMO pressure) and parasitaemia monitored over 96 h with SYBR Green I fluorescence (10 000 infected erythrocytes counted). * P
    Figure Legend Snippet: DFMO arrest and putrescine reversal associated to G1/S cell cycle control in intraerythrocytic P. falciparum parasites. In all instances, ■ refers to control parasites, ▼ refers to DFMO-treated parasites, and ● refers to putrescine-reversed parasites. ( a ) DFMO dose-response curves of asexual P . falciparum 3D7 proliferation over 96 h (initiated with ring-stage parasites, 1% haematocrit, 1% parasitaemia) at 37 °C in the absence or presence of 2 mM putrescine. Proliferation is expressed relative to untreated controls, with data averaged from n = 6 biological replicates and shown ± S.E. ( b ) Synchronised P . falciparum 3D7 cultures were treated with DFMO alone (IC 90 ) or with putrescine (2 mM, after 24 h DFMO pressure) and parasitaemia monitored over 96 h with SYBR Green I fluorescence (10 000 infected erythrocytes counted). * P

    Techniques Used: SYBR Green Assay, Fluorescence, Infection

    57) Product Images from "Hippocampal Telomerase Is Involved in the Modulation of Depressive Behaviors"

    Article Title: Hippocampal Telomerase Is Involved in the Modulation of Depressive Behaviors

    Journal: The Journal of Neuroscience

    doi: 10.1523/JNEUROSCI.0805-11.2011

    Implication of hippocampal telomerase alterations in depressive behaviors. A , Representative SYBR Green I-stained gel visualizing telomerase activity by TRAP products in the hippocampus of mice exposed to CMS for 21 d. Negative control 1, Taq enzyme-free control; Negative control 2, cell-free control; Positive control, cell line expressing a high level of telomerase; Heat-inactivated control, positive control sample incubated at 85°C for 10 min before the assay. Arrowheads indicate the key bands. B , Immunoblots showing protein level of TERT in the hippocampus of mice ( n = 3). C , Telomerase activity in the hippocampus of mice ( n = 3). In B and C , the mice were exposed to CMS for 49 d and treated with fluoxetine or vehicle during the last 28 d of CMS. D , Left, Representative DG area with Ad-mTERT-GFP transfection. Middle, Higher magnification of the boxed area. Right, RT-PCR analysis showing mRNA levels of mTERT in the hippocampus ( n = 3). E , Immobility time in the TST in mice treated with Ad-mTERT-GFP or Ad-GFP at 3 or 28 d after intrahippocampal microinjection ( n = 10–12). F–I , Effects of intrahippocampal Ad-mTERT-GFP transfection on CMS-induced behavioral modifications. Ad-mTERT-GFP or Ad-GFP was delivered into the bilateral DG of mice by microinjection; 4 d later, the mice were exposed to CMS for 21 d, and their body weight gain ( F ), immobility time in the TST ( G ) and FST ( H ), and sucrose preference ( I ) were examined on the next day ( n = 13–16). Data are mean ± SEM. * p
    Figure Legend Snippet: Implication of hippocampal telomerase alterations in depressive behaviors. A , Representative SYBR Green I-stained gel visualizing telomerase activity by TRAP products in the hippocampus of mice exposed to CMS for 21 d. Negative control 1, Taq enzyme-free control; Negative control 2, cell-free control; Positive control, cell line expressing a high level of telomerase; Heat-inactivated control, positive control sample incubated at 85°C for 10 min before the assay. Arrowheads indicate the key bands. B , Immunoblots showing protein level of TERT in the hippocampus of mice ( n = 3). C , Telomerase activity in the hippocampus of mice ( n = 3). In B and C , the mice were exposed to CMS for 49 d and treated with fluoxetine or vehicle during the last 28 d of CMS. D , Left, Representative DG area with Ad-mTERT-GFP transfection. Middle, Higher magnification of the boxed area. Right, RT-PCR analysis showing mRNA levels of mTERT in the hippocampus ( n = 3). E , Immobility time in the TST in mice treated with Ad-mTERT-GFP or Ad-GFP at 3 or 28 d after intrahippocampal microinjection ( n = 10–12). F–I , Effects of intrahippocampal Ad-mTERT-GFP transfection on CMS-induced behavioral modifications. Ad-mTERT-GFP or Ad-GFP was delivered into the bilateral DG of mice by microinjection; 4 d later, the mice were exposed to CMS for 21 d, and their body weight gain ( F ), immobility time in the TST ( G ) and FST ( H ), and sucrose preference ( I ) were examined on the next day ( n = 13–16). Data are mean ± SEM. * p

    Techniques Used: SYBR Green Assay, Staining, Activity Assay, Mouse Assay, Negative Control, Positive Control, Expressing, Incubation, Western Blot, Transfection, Reverse Transcription Polymerase Chain Reaction

    AZT inhibits telomerase activity and produces depressive-like phenotype. A , Representative SYBR Green I-stained gel visualizing TRAP products (left) and telomerase activity (right) in the hippocampus of mice treated with 25, 50, or 100 mg/kg AZT for 7 d ( n = 3). Negative control, Taq enzyme-free control; positive control, cell line expressing high levels of telomerase. B , Time scan of fluorescence intensity of samples showing DNA polymerase activity in the hippocampus of mice treated with 100 mg/kg AZT or vehicle for 7 d ( n = 3). KF, Klenow fragment (DNA polymerase I), a positive control; KF−, without KF, a negative control. C–H , Effects of AZT on behavioral phenotype. Shown are the immobility time in the TST ( C ) and FST ( D ) and percentage sucrose preference ( E ) of the mice treated with AZT (100 mg/kg, i.p.) for 14 d at the time indicated after withdrawal ( n = 10) and the immobility time in the TST ( F ) and FST ( G ) and percentage sucrose preference ( H ) of the mice treated with intrahippocampal AZT microinjection (0.7 μmol, 2 μl) at the time indicated after microinjections ( n = 9–11). Data are mean ± SEM. * p
    Figure Legend Snippet: AZT inhibits telomerase activity and produces depressive-like phenotype. A , Representative SYBR Green I-stained gel visualizing TRAP products (left) and telomerase activity (right) in the hippocampus of mice treated with 25, 50, or 100 mg/kg AZT for 7 d ( n = 3). Negative control, Taq enzyme-free control; positive control, cell line expressing high levels of telomerase. B , Time scan of fluorescence intensity of samples showing DNA polymerase activity in the hippocampus of mice treated with 100 mg/kg AZT or vehicle for 7 d ( n = 3). KF, Klenow fragment (DNA polymerase I), a positive control; KF−, without KF, a negative control. C–H , Effects of AZT on behavioral phenotype. Shown are the immobility time in the TST ( C ) and FST ( D ) and percentage sucrose preference ( E ) of the mice treated with AZT (100 mg/kg, i.p.) for 14 d at the time indicated after withdrawal ( n = 10) and the immobility time in the TST ( F ) and FST ( G ) and percentage sucrose preference ( H ) of the mice treated with intrahippocampal AZT microinjection (0.7 μmol, 2 μl) at the time indicated after microinjections ( n = 9–11). Data are mean ± SEM. * p

    Techniques Used: Activity Assay, SYBR Green Assay, Staining, Mouse Assay, Negative Control, Positive Control, Expressing, Fluorescence

    58) Product Images from "Simple flow cytometric detection of haemozoin containing leukocytes and erythrocytes for research on diagnosis, immunology and drug sensitivity testing"

    Article Title: Simple flow cytometric detection of haemozoin containing leukocytes and erythrocytes for research on diagnosis, immunology and drug sensitivity testing

    Journal: Malaria Journal

    doi: 10.1186/1475-2875-10-74

    Degree of depolarization corresponds to different developmental stages of P. berghei ANKA . Blood from a C57BL/6 mouse with a parasitaemia of 9.7% as determined by microscopy. Top plot shows several populations with decreasing degree of depolarization (A, B, C). Gate D shows low or non-depolarizing events. The four histograms show the number of events (n) and their fluorescence intensity (x-axis) of SYBR Green I, representing the DNA content. Gate D contains a population of 7.24% (arrow), likely including ring-forms with little Hz and thus very low or no depolarization.
    Figure Legend Snippet: Degree of depolarization corresponds to different developmental stages of P. berghei ANKA . Blood from a C57BL/6 mouse with a parasitaemia of 9.7% as determined by microscopy. Top plot shows several populations with decreasing degree of depolarization (A, B, C). Gate D shows low or non-depolarizing events. The four histograms show the number of events (n) and their fluorescence intensity (x-axis) of SYBR Green I, representing the DNA content. Gate D contains a population of 7.24% (arrow), likely including ring-forms with little Hz and thus very low or no depolarization.

    Techniques Used: Microscopy, Fluorescence, SYBR Green Assay

    59) Product Images from "DNA methyltransferase 3b preferentially associates with condensed chromatin"

    Article Title: DNA methyltransferase 3b preferentially associates with condensed chromatin

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkq870

    Dnmt3b is enriched in histone H1-containing chromatin fractions. ( A ) Dnmt3b, Dnmt3a2, Dnmt3a, Dnmt3L and Oct3/4 expression during ES cell differentiation. Equal amounts of protein (10 µg) from undifferentiated ES cells (Day 0) and differentiated ES cells (from Day 1 to 10) were analyzed by immunoblot using the indicated antibodies. ( B ) Subcellular distribution of Dnmt3b, Dnmt3a2 and Dnmt3L. ES cells were sequentially extracted to obtain cytoplasmic, chromatin and nuclear matrix fractions. Equal amounts of fractionated protein (10 µg) were separated by SDS–PAGE and then analyzed by immunoblot using the indicated antibodies. Fractionation efficiency was confirmed by immunoblot analysis using anti-PCNA (cytoplasm) and anti-lamin B1 (nuclear matrix) antibodies, and by Coomassie Brilliant Blue (CBB) staining of core histones (chromatin). ( C ) Schematic representation of chromatin fractionation. Isolated ES cell nuclei (N) were digested with 5, 20 or 80 U of MNase/mg DNA. Digested nuclei were fractionated into three fractions: transcriptionally active (S1), transcriptionally inactive (S2) and nuclear matrix-containing (P). ( D ) DNA samples from nuclear fractions N, S1, S2 and P. Equivalent amounts of DNA from each fraction were separated by 1.5% agarose gel electrophoresis in 1× TAE followed by SYBR-Green I staining. ( E ) Nuclear aliquots corresponding to 600 ng of DNA from N, S1, S2 and P fractions were separated by SDS–PAGE and then analyzed by immunoblot using the indicated antibodies.
    Figure Legend Snippet: Dnmt3b is enriched in histone H1-containing chromatin fractions. ( A ) Dnmt3b, Dnmt3a2, Dnmt3a, Dnmt3L and Oct3/4 expression during ES cell differentiation. Equal amounts of protein (10 µg) from undifferentiated ES cells (Day 0) and differentiated ES cells (from Day 1 to 10) were analyzed by immunoblot using the indicated antibodies. ( B ) Subcellular distribution of Dnmt3b, Dnmt3a2 and Dnmt3L. ES cells were sequentially extracted to obtain cytoplasmic, chromatin and nuclear matrix fractions. Equal amounts of fractionated protein (10 µg) were separated by SDS–PAGE and then analyzed by immunoblot using the indicated antibodies. Fractionation efficiency was confirmed by immunoblot analysis using anti-PCNA (cytoplasm) and anti-lamin B1 (nuclear matrix) antibodies, and by Coomassie Brilliant Blue (CBB) staining of core histones (chromatin). ( C ) Schematic representation of chromatin fractionation. Isolated ES cell nuclei (N) were digested with 5, 20 or 80 U of MNase/mg DNA. Digested nuclei were fractionated into three fractions: transcriptionally active (S1), transcriptionally inactive (S2) and nuclear matrix-containing (P). ( D ) DNA samples from nuclear fractions N, S1, S2 and P. Equivalent amounts of DNA from each fraction were separated by 1.5% agarose gel electrophoresis in 1× TAE followed by SYBR-Green I staining. ( E ) Nuclear aliquots corresponding to 600 ng of DNA from N, S1, S2 and P fractions were separated by SDS–PAGE and then analyzed by immunoblot using the indicated antibodies.

    Techniques Used: Expressing, Cell Differentiation, SDS Page, Fractionation, Staining, Isolation, Agarose Gel Electrophoresis, SYBR Green Assay

    Dnmt3b associates with nuclease-resistant heterochromatin and the association requires higher-order chromatin structure. ( A ) DNA from nuclei digested with 5 or 80 U MNase/mg DNA. Equivalent amounts of DNA were separated by 1.5% agarose gel electrophoresis in 1× TAE and then visualized with SYBR-green I. ( B ) Detection of chromatin in FLAG-Dnmt3b immunoprecipitates. Nuclear extracts were digested with 5 or 80 U MNase/mg DNA, and subjected to immunoprecipitation using an anti-FLAG antibody in the presence of 75 mM NaCl. FLAG-Dnmt3b immunoprecipitates were analyzed by immunoblot using an anti-histone H3 antibody to confirm the interaction between Dnmt3b and chromatin. Input represents 4% of the nuclear extract used in the assay. ( C ) Dnmt3b co-sedimented with histone H1-containing poly-nucleosomes on a sucrose gradient. S2 fractions were subjected to 10–30% (w/v) sucrose gradient sedimentation. Equivalent aliquots of each DNA fraction and protein sample were analyzed by agarose gel electrophoresis and immunoblot using the indicated antibodies, respectively. Core histones in each fraction were visualized by CBB staining.
    Figure Legend Snippet: Dnmt3b associates with nuclease-resistant heterochromatin and the association requires higher-order chromatin structure. ( A ) DNA from nuclei digested with 5 or 80 U MNase/mg DNA. Equivalent amounts of DNA were separated by 1.5% agarose gel electrophoresis in 1× TAE and then visualized with SYBR-green I. ( B ) Detection of chromatin in FLAG-Dnmt3b immunoprecipitates. Nuclear extracts were digested with 5 or 80 U MNase/mg DNA, and subjected to immunoprecipitation using an anti-FLAG antibody in the presence of 75 mM NaCl. FLAG-Dnmt3b immunoprecipitates were analyzed by immunoblot using an anti-histone H3 antibody to confirm the interaction between Dnmt3b and chromatin. Input represents 4% of the nuclear extract used in the assay. ( C ) Dnmt3b co-sedimented with histone H1-containing poly-nucleosomes on a sucrose gradient. S2 fractions were subjected to 10–30% (w/v) sucrose gradient sedimentation. Equivalent aliquots of each DNA fraction and protein sample were analyzed by agarose gel electrophoresis and immunoblot using the indicated antibodies, respectively. Core histones in each fraction were visualized by CBB staining.

    Techniques Used: Agarose Gel Electrophoresis, SYBR Green Assay, Immunoprecipitation, Sedimentation, Staining

    60) Product Images from "A kinetic-based sigmoidal model for the polymerase chain reaction and its application to high-capacity absolute quantitative real-time PCR"

    Article Title: A kinetic-based sigmoidal model for the polymerase chain reaction and its application to high-capacity absolute quantitative real-time PCR

    Journal: BMC Biotechnology

    doi: 10.1186/1472-6750-8-47

    LRE analysis of eleven cDNA targets amplified with five reaction formulations . Summaries for LRE analysis of a total of 134 amplification profiles (each profile an average of four replicate amplification reactions) generated from eleven cDNA targets quantified using five reaction formulations (see Figure 2 and additional file 3 for additional details). (A) Average linear regression correlation coefficients (r 2 ). (B) Average E max . (C) Average ΔE. Note that ΔE is dependent on the optics of the assay and are thus not directly comparable across different formulations and gain settings. (D) Average F 0 derived from the average of F 0 values encompassed by the LRE window (see Figure 4 for details). QT : QuantiTect, DyNa : DyNAmo; FV : FullVelocity, SG : SYBR Green I, SD : standard deviation, n : the number of runs, na : not applicable (only one run conducted), nd : not determined.
    Figure Legend Snippet: LRE analysis of eleven cDNA targets amplified with five reaction formulations . Summaries for LRE analysis of a total of 134 amplification profiles (each profile an average of four replicate amplification reactions) generated from eleven cDNA targets quantified using five reaction formulations (see Figure 2 and additional file 3 for additional details). (A) Average linear regression correlation coefficients (r 2 ). (B) Average E max . (C) Average ΔE. Note that ΔE is dependent on the optics of the assay and are thus not directly comparable across different formulations and gain settings. (D) Average F 0 derived from the average of F 0 values encompassed by the LRE window (see Figure 4 for details). QT : QuantiTect, DyNa : DyNAmo; FV : FullVelocity, SG : SYBR Green I, SD : standard deviation, n : the number of runs, na : not applicable (only one run conducted), nd : not determined.

    Techniques Used: Amplification, Generated, Derivative Assay, SYBR Green Assay, Standard Deviation

    Assessing the impact of SYBR Green I quantity on real-time amplification profiles . Replicate amplification reactions were prepared using three commercial enzyme formulations supplemented with increasing amounts of SYBR Green I. Each amplification reaction contained 100 femtograms of lambda gDNA (1,876 genomes) and 500 μM of the primers K7B and K12. The gain setting for each run was adjusted to ensure that reaction fluorescence remained below the saturation level of the photomultiplier tube (about 40,000 FU). (A) , (C) and (E) Screen shots of amplification profiles generated by the Stratagene MxPro qPCR software. This reveals that fluorescence intensity is dependent on SYBR Green I quantity, reflected by the large increase in the height of the amplification profiles as SYBR Green I quantity is increased. (B) , (D) and (F) LRE plot of each respective amplification profile. This reveals a linear domain corresponding to the central region of each amplification profile, confirming the mathematical prediction that amplification efficiency is linearly coupled to amplicon quantity. A consecutive group of points was selected (designated by red circles) for linear regression analysis (referred to as
    Figure Legend Snippet: Assessing the impact of SYBR Green I quantity on real-time amplification profiles . Replicate amplification reactions were prepared using three commercial enzyme formulations supplemented with increasing amounts of SYBR Green I. Each amplification reaction contained 100 femtograms of lambda gDNA (1,876 genomes) and 500 μM of the primers K7B and K12. The gain setting for each run was adjusted to ensure that reaction fluorescence remained below the saturation level of the photomultiplier tube (about 40,000 FU). (A) , (C) and (E) Screen shots of amplification profiles generated by the Stratagene MxPro qPCR software. This reveals that fluorescence intensity is dependent on SYBR Green I quantity, reflected by the large increase in the height of the amplification profiles as SYBR Green I quantity is increased. (B) , (D) and (F) LRE plot of each respective amplification profile. This reveals a linear domain corresponding to the central region of each amplification profile, confirming the mathematical prediction that amplification efficiency is linearly coupled to amplicon quantity. A consecutive group of points was selected (designated by red circles) for linear regression analysis (referred to as "LRE analysis"), generating estimates for E max (intercept) and ΔE (slope), from which F max was calculated using equation 4 (see Figure 1 for additional details). Details as to how the boundaries of the linear region were determined are described later in the study. [SG] : quantity of supplementary SYBR Green I, r2 : linear regression correlation coefficient.

    Techniques Used: SYBR Green Assay, Amplification, Fluorescence, Generated, Real-time Polymerase Chain Reaction, Software

    Optimizing LRE window size via recursion . This data is taken from the 0.5X SYBR Green I DyNAmo amplification profile (Figure 2); however, all the amplification profiles in Figure 2 generate similar results. The recursive analysis starts by assigning a small LRE window (designated by red circles) in the lower region of the amplification profile and conducting linear regression analysis using the points within this preliminary LRE window (blue highlight). F C readings are then converted to F 0 using equation 6 and the average determined for F 0 values encompassed by the LRE window (green highlight). Conformity of the F 0 value generated by the cycle immediate following the last cycle of the LRE window is then assessed (bold), based upon the percent difference with the LRE window F 0 average (%Diff, yellow highlight). If this difference is less than a specified threshold, the LRE window is expanded to include the next cycle and the analysis repeated. This recursive process is continued until a nonconforming cycle is encountered as defined by the threshold. Note that for illustrative purposes, the LRE window in this figure has been expanded beyond the threshold used in this study (7.5%). Although the LRE line generated within the LRE plot can be used to assess conformity, examining the resulting F 0 values provides a more sensitive and objective methodology, as is visually illustrated by the F 0 plots.
    Figure Legend Snippet: Optimizing LRE window size via recursion . This data is taken from the 0.5X SYBR Green I DyNAmo amplification profile (Figure 2); however, all the amplification profiles in Figure 2 generate similar results. The recursive analysis starts by assigning a small LRE window (designated by red circles) in the lower region of the amplification profile and conducting linear regression analysis using the points within this preliminary LRE window (blue highlight). F C readings are then converted to F 0 using equation 6 and the average determined for F 0 values encompassed by the LRE window (green highlight). Conformity of the F 0 value generated by the cycle immediate following the last cycle of the LRE window is then assessed (bold), based upon the percent difference with the LRE window F 0 average (%Diff, yellow highlight). If this difference is less than a specified threshold, the LRE window is expanded to include the next cycle and the analysis repeated. This recursive process is continued until a nonconforming cycle is encountered as defined by the threshold. Note that for illustrative purposes, the LRE window in this figure has been expanded beyond the threshold used in this study (7.5%). Although the LRE line generated within the LRE plot can be used to assess conformity, examining the resulting F 0 values provides a more sensitive and objective methodology, as is visually illustrated by the F 0 plots.

    Techniques Used: SYBR Green Assay, Amplification, Generated

    Optical calibrations of five reaction formulations supplemented with various quantities of SYBR Green I . Spreadsheet summary of the lambda gDNA (primer pair K7B-K12) optical calibrations conducted for the five reaction formulations used for LRE quantification in this study. Based on data presented in Figure 2, these formulations were designed to assess the impact of SYBR Green I quantity, the highest supplemented quantity (2.0X) estimated to be > 10X than that present in these three commercial formulations. Each row of values was derived from an individual amplification run (see additional file 3 for more details). (A) QuantiTect 0X SYBR Green I. (B) QuantiTect 0.2X SYBR Green I. (C) DyNAmo 0.5X SYBR Green I. (D) DyNAmo 1.5X SYBR Green I. (E) FullVelocity 2.0X SYBR Green I. Lam : lambda gDNA quantity in femtograms, QT : QuantiTect, DyNa : DyNAmo; FV : FullVelocity, SG : SYBR Green I, SD : standard deviation, CV : coefficient of variation (SD/Average × 100%).
    Figure Legend Snippet: Optical calibrations of five reaction formulations supplemented with various quantities of SYBR Green I . Spreadsheet summary of the lambda gDNA (primer pair K7B-K12) optical calibrations conducted for the five reaction formulations used for LRE quantification in this study. Based on data presented in Figure 2, these formulations were designed to assess the impact of SYBR Green I quantity, the highest supplemented quantity (2.0X) estimated to be > 10X than that present in these three commercial formulations. Each row of values was derived from an individual amplification run (see additional file 3 for more details). (A) QuantiTect 0X SYBR Green I. (B) QuantiTect 0.2X SYBR Green I. (C) DyNAmo 0.5X SYBR Green I. (D) DyNAmo 1.5X SYBR Green I. (E) FullVelocity 2.0X SYBR Green I. Lam : lambda gDNA quantity in femtograms, QT : QuantiTect, DyNa : DyNAmo; FV : FullVelocity, SG : SYBR Green I, SD : standard deviation, CV : coefficient of variation (SD/Average × 100%).

    Techniques Used: SYBR Green Assay, Derivative Assay, Amplification, Laser Capture Microdissection, Standard Deviation

    61) Product Images from "Investigation of Förster Resonance Energy Transfer (FRET) and Competition of Fluorescent Dyes on DNA Microparticles"

    Article Title: Investigation of Förster Resonance Energy Transfer (FRET) and Competition of Fluorescent Dyes on DNA Microparticles

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms16047738

    Alteration of the FRET dependent on the staining methods. The dsDNA balls were stained with the green and orange dyes simultaneously or in series. In the serial staining, SYTOX Orange was added after SYBR green staining ( a ) SYBR Green I was used as the green fluorescent dye; ( b ) SYBR Green II was used as the green fluorescent dye; and ( c ) Decrease in the green fluorescence intensity depending on the staining methods.
    Figure Legend Snippet: Alteration of the FRET dependent on the staining methods. The dsDNA balls were stained with the green and orange dyes simultaneously or in series. In the serial staining, SYTOX Orange was added after SYBR green staining ( a ) SYBR Green I was used as the green fluorescent dye; ( b ) SYBR Green II was used as the green fluorescent dye; and ( c ) Decrease in the green fluorescence intensity depending on the staining methods.

    Techniques Used: Staining, SYBR Green Assay, Fluorescence

    62) Product Images from "SNP Genotyping by Multiplexed Solid-Phase Amplification and Fluorescent Minisequencing"

    Article Title: SNP Genotyping by Multiplexed Solid-Phase Amplification and Fluorescent Minisequencing

    Journal: Genome Research

    doi: 10.1101/gr.205001

    Single-nucleotide polymorphism (SNP) minisequencing. ( A ) Diagram of the components and products of solid-phase amplification and SNP minisequencing using 5HT 2A -specific primers. The polymorphic site (C/T) is shown in parentheses. The fluorescently labeled nucleotides (G* or A*) are depicted following Bbv I digestion and nucleotide incorporation. ( B ) Fluorescent image of 10% acrylamide gel with single-color FAM-ddNTP minisequencing of 5-HT 2A solid-phase PCR product. ( C ) Sybr green I staining of same gel from panel B . ( D ) Four-color minisequencing of 5HT 2A locus.
    Figure Legend Snippet: Single-nucleotide polymorphism (SNP) minisequencing. ( A ) Diagram of the components and products of solid-phase amplification and SNP minisequencing using 5HT 2A -specific primers. The polymorphic site (C/T) is shown in parentheses. The fluorescently labeled nucleotides (G* or A*) are depicted following Bbv I digestion and nucleotide incorporation. ( B ) Fluorescent image of 10% acrylamide gel with single-color FAM-ddNTP minisequencing of 5-HT 2A solid-phase PCR product. ( C ) Sybr green I staining of same gel from panel B . ( D ) Four-color minisequencing of 5HT 2A locus.

    Techniques Used: Amplification, Labeling, Acrylamide Gel Assay, Polymerase Chain Reaction, SYBR Green Assay, Staining

    63) Product Images from "An Optimized SYBR Green I/PI Assay for Rapid Viability Assessment and Antibiotic Susceptibility Testing for Borrelia burgdorferi"

    Article Title: An Optimized SYBR Green I/PI Assay for Rapid Viability Assessment and Antibiotic Susceptibility Testing for Borrelia burgdorferi

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0111809

    Representative images of B. burgdorferi culture (7 day old), observed with fluorescence microscopy equipped with Spot slider color camera using LIVE/DEAD BacLight stain (A), SYBR Green I/PI stain (B), and FDA stain (C). Antibiotic-treated B. burgdorferi biofilm (9 day old) was stained by SYBR Green I/PI (D). Sytox Green/Hoechst 33342 stained B. burgdorferi images were recorded by the ORCA-R 2 high resolution camera (E, merged from images visualized by DAPI, FITC and TRITC filters) and by Spot slider color camera with triple filter (F).
    Figure Legend Snippet: Representative images of B. burgdorferi culture (7 day old), observed with fluorescence microscopy equipped with Spot slider color camera using LIVE/DEAD BacLight stain (A), SYBR Green I/PI stain (B), and FDA stain (C). Antibiotic-treated B. burgdorferi biofilm (9 day old) was stained by SYBR Green I/PI (D). Sytox Green/Hoechst 33342 stained B. burgdorferi images were recorded by the ORCA-R 2 high resolution camera (E, merged from images visualized by DAPI, FITC and TRITC filters) and by Spot slider color camera with triple filter (F).

    Techniques Used: Fluorescence, Microscopy, Staining, SYBR Green Assay

    Linear relationship between the B. burgdorferi viability and Green/Red fluorescence ratio of the SYBR Green I/PI assay. (A) The linear relationship between the number of spirochetes and Green/Red fluorescence ratio. (B) Emission spectra of suspensions of various proportions of live and isopropyl alcohol-killed B. burgdorferi were obtained, and the Green/Red fluorescence ratios were calculated for each proportion of live/dead cells. The line is a least-square fit of the relationship between percentage of live bacteria and Green/Red fluorescence ratio.
    Figure Legend Snippet: Linear relationship between the B. burgdorferi viability and Green/Red fluorescence ratio of the SYBR Green I/PI assay. (A) The linear relationship between the number of spirochetes and Green/Red fluorescence ratio. (B) Emission spectra of suspensions of various proportions of live and isopropyl alcohol-killed B. burgdorferi were obtained, and the Green/Red fluorescence ratios were calculated for each proportion of live/dead cells. The line is a least-square fit of the relationship between percentage of live bacteria and Green/Red fluorescence ratio.

    Techniques Used: Fluorescence, SYBR Green Assay

    The Green/Red fluorescence ratios of B. burgdorferi biofilm measured by the SYBR Green I/PI assay at different culture times. The top panel shows the increase in Green/Red fluorescence ratios for biofilm growth over time, whereas the lower panel shows the corresponding microscopic images at different time points. All assays were run in triplicate.
    Figure Legend Snippet: The Green/Red fluorescence ratios of B. burgdorferi biofilm measured by the SYBR Green I/PI assay at different culture times. The top panel shows the increase in Green/Red fluorescence ratios for biofilm growth over time, whereas the lower panel shows the corresponding microscopic images at different time points. All assays were run in triplicate.

    Techniques Used: Fluorescence, SYBR Green Assay

    64) Product Images from "A molecular, phylogenetic and functional study of the dADAR mRNA truncated isoform during Drosophila embryonic development reveals an editing-independent function"

    Article Title: A molecular, phylogenetic and functional study of the dADAR mRNA truncated isoform during Drosophila embryonic development reveals an editing-independent function

    Journal: Open journal of animal sciences

    doi: 10.4236/ojas.2013.34A2003

    Semi-quantitative RT-PCR for dADAR mRNA isoform relative abundance in D. melanogaster. Orientation diagram (a) shows positions of isoform-specific PCR primers. Gel electrophoresis of PCR products (b) shows relatively high levels of full-length isoform (Ex-7) in comparison to truncated isoform (In-6) in ethidium bromide stained gel. Quantification of band intensities in gels stained with SYBR green I is shown in panel (c). Constitutively expressed rp -49 mRNA was included as a reference against which the other band intensities were compared.
    Figure Legend Snippet: Semi-quantitative RT-PCR for dADAR mRNA isoform relative abundance in D. melanogaster. Orientation diagram (a) shows positions of isoform-specific PCR primers. Gel electrophoresis of PCR products (b) shows relatively high levels of full-length isoform (Ex-7) in comparison to truncated isoform (In-6) in ethidium bromide stained gel. Quantification of band intensities in gels stained with SYBR green I is shown in panel (c). Constitutively expressed rp -49 mRNA was included as a reference against which the other band intensities were compared.

    Techniques Used: Quantitative RT-PCR, Polymerase Chain Reaction, Nucleic Acid Electrophoresis, Staining, SYBR Green Assay

    65) Product Images from "iFRET: An Improved Fluorescence System for DNA-Melting Analysis"

    Article Title: iFRET: An Improved Fluorescence System for DNA-Melting Analysis

    Journal: Genome Research

    doi: 10.1101/gr.297202

    Comparision of signal intensity trends for Sybr Green I, Fluorescence resonance energy transfer (FRET), and induced (i)FRET. Melting curves are shown for the three indicated assay formats. Fluorescence measurements were recorded at the empirically determined emission maxima for respective dyes: for SYBR Green I detection: 541–545 nm, for FRET detection (FAM): 541–545 nm, and for iFRET detection (ROX): 611–615 nm.
    Figure Legend Snippet: Comparision of signal intensity trends for Sybr Green I, Fluorescence resonance energy transfer (FRET), and induced (i)FRET. Melting curves are shown for the three indicated assay formats. Fluorescence measurements were recorded at the empirically determined emission maxima for respective dyes: for SYBR Green I detection: 541–545 nm, for FRET detection (FAM): 541–545 nm, and for iFRET detection (ROX): 611–615 nm.

    Techniques Used: SYBR Green Assay, Fluorescence, Förster Resonance Energy Transfer

    66) Product Images from "Counting Viruses and Bacteria in Photosynthetic Microbial Mats"

    Article Title: Counting Viruses and Bacteria in Photosynthetic Microbial Mats

    Journal: Applied and Environmental Microbiology

    doi: 10.1128/AEM.02863-14

    Cytogram of viruses from photosynthetic microbial mat samples using flow cytometry after staining with nucleic acid-specific dye SYBR green I (A) and from control sample without viruses (B). Green fluorescence (V1 and V2) allows the distinction of two virus clusters.
    Figure Legend Snippet: Cytogram of viruses from photosynthetic microbial mat samples using flow cytometry after staining with nucleic acid-specific dye SYBR green I (A) and from control sample without viruses (B). Green fluorescence (V1 and V2) allows the distinction of two virus clusters.

    Techniques Used: Flow Cytometry, Cytometry, Staining, SYBR Green Assay, Fluorescence

    67) Product Images from "Identification of novel activity against Borrelia burgdorferi persisters using an FDA approved drug library"

    Article Title: Identification of novel activity against Borrelia burgdorferi persisters using an FDA approved drug library

    Journal: Emerging Microbes & Infections

    doi: 10.1038/emi.2014.53

    Representative images of stationary-phase B. burgdorferi strain B31 treated with different antibiotics (50 µM) followed by staining in the SYBR Green I/PI assay (×400 magnification). ( A ) Drug-free control, ( B ) Doxycycline, ( C ) Amoxicillin, ( D ) Daptomycin, ( E ) Cefoperazone, ( F ) Clofazimine, ( G ) Carbomycin, ( H ) Cefotiam, and ( I ) Tetracycline.
    Figure Legend Snippet: Representative images of stationary-phase B. burgdorferi strain B31 treated with different antibiotics (50 µM) followed by staining in the SYBR Green I/PI assay (×400 magnification). ( A ) Drug-free control, ( B ) Doxycycline, ( C ) Amoxicillin, ( D ) Daptomycin, ( E ) Cefoperazone, ( F ) Clofazimine, ( G ) Carbomycin, ( H ) Cefotiam, and ( I ) Tetracycline.

    Techniques Used: Staining, SYBR Green Assay

    ( A ) Growth curve of B. burgdorferi strain B31 in vitro . ( B ) Representative images of the log phase (3-day culture) and stationary phase of B. burgdorferi B31 strain (7-day culture), observed with fluorescent microscopy using the SYBR Green I/PI stain (×400 magnification). The arrows indicate multiple morphological forms of B. burgdorferi in stationary phase.
    Figure Legend Snippet: ( A ) Growth curve of B. burgdorferi strain B31 in vitro . ( B ) Representative images of the log phase (3-day culture) and stationary phase of B. burgdorferi B31 strain (7-day culture), observed with fluorescent microscopy using the SYBR Green I/PI stain (×400 magnification). The arrows indicate multiple morphological forms of B. burgdorferi in stationary phase.

    Techniques Used: In Vitro, Microscopy, SYBR Green Assay, Staining

    Susceptibility of log phase (3 days) and stationary-phase (7 days) B. burgdorferi to 50 µM drugs after a 5-day treatment. The percentages of residual live cells were determined using the SYBR Green I/PI assay.
    Figure Legend Snippet: Susceptibility of log phase (3 days) and stationary-phase (7 days) B. burgdorferi to 50 µM drugs after a 5-day treatment. The percentages of residual live cells were determined using the SYBR Green I/PI assay.

    Techniques Used: SYBR Green Assay

    68) Product Images from "Fluorophore-Labeled Primers Improve the Sensitivity, Versatility, and Normalization of Denaturing Gradient Gel Electrophoresis"

    Article Title: Fluorophore-Labeled Primers Improve the Sensitivity, Versatility, and Normalization of Denaturing Gradient Gel Electrophoresis

    Journal: Applied and Environmental Microbiology

    doi: 10.1128/AEM.71.8.4893-4896.2005

    Impact of fluorophore incorporation on DGGE fingerprints. This SYBR Green I-stained gel contains PCR products from three soils with and without fluorophore labels. An asterisk indicates the position of two gradient-sensitive bands in the S14 fingerprints
    Figure Legend Snippet: Impact of fluorophore incorporation on DGGE fingerprints. This SYBR Green I-stained gel contains PCR products from three soils with and without fluorophore labels. An asterisk indicates the position of two gradient-sensitive bands in the S14 fingerprints

    Techniques Used: Denaturing Gradient Gel Electrophoresis, SYBR Green Assay, Staining, Polymerase Chain Reaction

    69) Product Images from "Saccharomyces cerevisiae HMO1 interacts with TFIID and participates in start site selection by RNA polymerase II"

    Article Title: Saccharomyces cerevisiae HMO1 interacts with TFIID and participates in start site selection by RNA polymerase II

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkm1068

    In vivo association of TAF1 with RPG promoters. ( A ) In vivo binding of TAF1 to the promoter region of RPS5 , RPS31 , RPL10 , RPL3 , RPL27B , RPL23B and RPL32 was analyzed using ChIP assays. Yeast strains were grown in YPD medium to mid-log phase at 25°C. Cross-linked chromatin from HMO1 or Δ hmo1 strains expressing TAP-tagged TAF1, with or without the TAF N-terminal domain (TAND; 2–186 aa), as indicated, was prepared and precipitated with either IgG-Sepharose 6 FastFlow (IgG; +) or Sepharose 6 FastFlow (IgG; −, negative control) beads. After reversal of cross-linking, PCR was performed to test for the presence of DNA corresponding to the promoter regions of the indicated genes. Each PCR reaction contained a second primer pair that amplifies a region (218 bp) of the POL1 ORF as an internal background control (asterisk) ( 27 ). The lower panel (input) shows the results of PCR conducted with the chromatin prior to precipitation. ( B ) Quantitation of the raw data shown in A. Signals corresponding to each band were quantified by an image analyzer after staining with SYBR Green I. The ratio of the precipitated signal (IP) to the input signal from each lysate (table at bottom) was calculated for all the indicated RPG promoters (top panel) as well as for the POL1 ORF (middle panel). Northern blot analysis was also conducted as described in Figure 3 A. The raw data (data not shown) were quantified and are presented graphically in the bottom panel. Values for each transcript were normalized to the maximum expression of that transcript.
    Figure Legend Snippet: In vivo association of TAF1 with RPG promoters. ( A ) In vivo binding of TAF1 to the promoter region of RPS5 , RPS31 , RPL10 , RPL3 , RPL27B , RPL23B and RPL32 was analyzed using ChIP assays. Yeast strains were grown in YPD medium to mid-log phase at 25°C. Cross-linked chromatin from HMO1 or Δ hmo1 strains expressing TAP-tagged TAF1, with or without the TAF N-terminal domain (TAND; 2–186 aa), as indicated, was prepared and precipitated with either IgG-Sepharose 6 FastFlow (IgG; +) or Sepharose 6 FastFlow (IgG; −, negative control) beads. After reversal of cross-linking, PCR was performed to test for the presence of DNA corresponding to the promoter regions of the indicated genes. Each PCR reaction contained a second primer pair that amplifies a region (218 bp) of the POL1 ORF as an internal background control (asterisk) ( 27 ). The lower panel (input) shows the results of PCR conducted with the chromatin prior to precipitation. ( B ) Quantitation of the raw data shown in A. Signals corresponding to each band were quantified by an image analyzer after staining with SYBR Green I. The ratio of the precipitated signal (IP) to the input signal from each lysate (table at bottom) was calculated for all the indicated RPG promoters (top panel) as well as for the POL1 ORF (middle panel). Northern blot analysis was also conducted as described in Figure 3 A. The raw data (data not shown) were quantified and are presented graphically in the bottom panel. Values for each transcript were normalized to the maximum expression of that transcript.

    Techniques Used: In Vivo, Binding Assay, Chromatin Immunoprecipitation, Expressing, Negative Control, Polymerase Chain Reaction, Quantitation Assay, Staining, SYBR Green Assay, Northern Blot

    70) Product Images from "Factors determining microbial colonization of liquid nitrogen storage tanks used for archiving biological samples"

    Article Title: Factors determining microbial colonization of liquid nitrogen storage tanks used for archiving biological samples

    Journal: Applied Microbiology and Biotechnology

    doi: 10.1007/s00253-019-10242-1

    Epifluorescence photomicrographs of SYBR Green I-stained samples and algae autoflourenscence. ( a ) bacterial cells detected in the negative control NC-B-2 containing 2 × 10 2 cells per ml, ( b ) bacterial cells detected in the ice sample B-4-2 containing 8 × 10 3 cells per ml, ( c ) eukaryotic cells of sample G-20-1, confirmed by PCR with Line1 primer, filaments of Cyanobacteria in sample I-30-3 ( d ) autofluorescence of microbial cells (excitation 425 nm, emission 630 nm) and ( e ) overlay with phase contrast. Sample code, institute-identity number-replicate; Scale bar, 5 μm
    Figure Legend Snippet: Epifluorescence photomicrographs of SYBR Green I-stained samples and algae autoflourenscence. ( a ) bacterial cells detected in the negative control NC-B-2 containing 2 × 10 2 cells per ml, ( b ) bacterial cells detected in the ice sample B-4-2 containing 8 × 10 3 cells per ml, ( c ) eukaryotic cells of sample G-20-1, confirmed by PCR with Line1 primer, filaments of Cyanobacteria in sample I-30-3 ( d ) autofluorescence of microbial cells (excitation 425 nm, emission 630 nm) and ( e ) overlay with phase contrast. Sample code, institute-identity number-replicate; Scale bar, 5 μm

    Techniques Used: SYBR Green Assay, Staining, Negative Control, Polymerase Chain Reaction

    71) Product Images from "Assembly of Regulatory Factors on rRNA and Ribosomal Protein Genes in Saccharomyces cerevisiae ▿ ▿ †"

    Article Title: Assembly of Regulatory Factors on rRNA and Ribosomal Protein Genes in Saccharomyces cerevisiae ▿ ▿ †

    Journal: Molecular and Cellular Biology

    doi: 10.1128/MCB.00876-07

    HMO1 associates throughout the 35S rRNA gene in a Pol I-dependent manner. (A) In vivo binding of HMO1 to various regions of the rRNA gene was analyzed by ChIP assays. The yeast strain expressing the TAP-tagged HMO1 (YKK74) was grown in YPD (yeast extract, peptone, dextrose) medium to mid-log phase at 30°C. Cross-linked chromatin from this strain was prepared and precipitated with either immunoglobulin G (IgG)-Sepharose 6 FastFlow (+IgG) or Sepharose 6 FastFlow (−IgG; negative control) beads. After the reversal of the cross-linking, PCR was performed to test for the presence of DNA corresponding to regions 1 to 14 (indicated schematically in panel B). The PCR products were separated by 5% polyacrylamide gel electrophoresis and stained with SYBR Green I. The top, middle, and bottom (input DNA) panels indicate the results of the PCR conducted for the chromatin after precipitation with or without IgG, or before precipitation, respectively. (B) The quantification of the raw data shown in panel A. The signals corresponding to each band were quantified by an image analyzer. The ratio of the IgG-precipitated signal (IP) to the input signal was calculated for the regions 1 to 14. 4/P and 14/T regions are overlapped with the promoter (P) and terminator (T) of the 35S RNA gene, respectively. (C) The effect of Δ hmo1 and/or Δ rpa135 on the growth of the yeast cells. The Δ rpa135 and Δ hmo1 Δ rpa135 strains carrying the plasmid pM5032 (encoding RPA135 ) were designated HMO1 RPA135 (YKK69) and Δ hmo1 RPA135 (YKK70), respectively. Similarly, the Δ rpa135 and Δ hmo1 Δ rpa135 strains carrying the helper plasmid pM5057 expressing 35S rRNA from the TEF1 promoter (schematically indicated in panel G) were designated HMO1 Δ rpa135 (YKK72) and Δ hmo1 Δ rpa135 (YKK100), respectively. These four strains were spotted onto YPD plates at three dilutions and grown at 30°C for 5 days. (D) The strains described in panel C were grown in YPD medium to mid-log phase at 30°C and total RNA was obtained from 1 × 10 7 cells, then separated by 1% agarose gel electrophoresis and stained with ethidium bromide. The positions of 25S and 18S rRNA are indicated on the left. The asterisk may correspond to 5S/5.8S rRNA and tRNA. Further analyses on this band were described in Fig. S1 in the supplemental material. (E) In vivo binding of HMO1 to the promoter (4/P) and terminator (14/T) regions of the chromosomal 35S rRNA gene were analyzed by ChIP assays in the HMO1 RPA135 (YTK8866, indicated as WT) and HMO1 Δ rpa135 (YTK8867, indicated as Δ) strains expressing the FLAG-tagged HMO1. The strains were grown in synthetic complete (SC) medium to mid-log phase at 25°C. The cross-linked chromatin was prepared and precipitated with (+) or without (−) anti-FLAG monoclonal antibodies. After reversal of the cross-linking, PCR was performed and analyzed as described in panel A to test for the presence of DNA corresponding to the regions of 4/P and 14/T, both of which are modified (thereby were not contained) in the helper plasmid as shown in panels B and G. Left panel (input) shows the results of the PCR conducted on the chromatin before precipitation. (F) The quantification of the raw data shown in panel E. The signals corresponding to each band were quantified by an image analyzer. The ratio of the precipitated signal (IP) to the input signal was calculated for the regions 4/P and 14/T. (G) A schematic diagram of the helper plasmid pM5057. The regions that were amplified by PCR in the ChIP assays (H and I) are indicated as P, T, and I. (H) In vivo binding of HMO1 to the promoter (P), terminator (T) and unrelated intermediate (I) regions of the 35S rRNA gene on the helper plasmid were analyzed by ChIP assays using the same DNA fractions (input and IP) as described in panel E. (I) The quantification of the raw data shown in panel H was performed as described in panel F. WT, wild type.
    Figure Legend Snippet: HMO1 associates throughout the 35S rRNA gene in a Pol I-dependent manner. (A) In vivo binding of HMO1 to various regions of the rRNA gene was analyzed by ChIP assays. The yeast strain expressing the TAP-tagged HMO1 (YKK74) was grown in YPD (yeast extract, peptone, dextrose) medium to mid-log phase at 30°C. Cross-linked chromatin from this strain was prepared and precipitated with either immunoglobulin G (IgG)-Sepharose 6 FastFlow (+IgG) or Sepharose 6 FastFlow (−IgG; negative control) beads. After the reversal of the cross-linking, PCR was performed to test for the presence of DNA corresponding to regions 1 to 14 (indicated schematically in panel B). The PCR products were separated by 5% polyacrylamide gel electrophoresis and stained with SYBR Green I. The top, middle, and bottom (input DNA) panels indicate the results of the PCR conducted for the chromatin after precipitation with or without IgG, or before precipitation, respectively. (B) The quantification of the raw data shown in panel A. The signals corresponding to each band were quantified by an image analyzer. The ratio of the IgG-precipitated signal (IP) to the input signal was calculated for the regions 1 to 14. 4/P and 14/T regions are overlapped with the promoter (P) and terminator (T) of the 35S RNA gene, respectively. (C) The effect of Δ hmo1 and/or Δ rpa135 on the growth of the yeast cells. The Δ rpa135 and Δ hmo1 Δ rpa135 strains carrying the plasmid pM5032 (encoding RPA135 ) were designated HMO1 RPA135 (YKK69) and Δ hmo1 RPA135 (YKK70), respectively. Similarly, the Δ rpa135 and Δ hmo1 Δ rpa135 strains carrying the helper plasmid pM5057 expressing 35S rRNA from the TEF1 promoter (schematically indicated in panel G) were designated HMO1 Δ rpa135 (YKK72) and Δ hmo1 Δ rpa135 (YKK100), respectively. These four strains were spotted onto YPD plates at three dilutions and grown at 30°C for 5 days. (D) The strains described in panel C were grown in YPD medium to mid-log phase at 30°C and total RNA was obtained from 1 × 10 7 cells, then separated by 1% agarose gel electrophoresis and stained with ethidium bromide. The positions of 25S and 18S rRNA are indicated on the left. The asterisk may correspond to 5S/5.8S rRNA and tRNA. Further analyses on this band were described in Fig. S1 in the supplemental material. (E) In vivo binding of HMO1 to the promoter (4/P) and terminator (14/T) regions of the chromosomal 35S rRNA gene were analyzed by ChIP assays in the HMO1 RPA135 (YTK8866, indicated as WT) and HMO1 Δ rpa135 (YTK8867, indicated as Δ) strains expressing the FLAG-tagged HMO1. The strains were grown in synthetic complete (SC) medium to mid-log phase at 25°C. The cross-linked chromatin was prepared and precipitated with (+) or without (−) anti-FLAG monoclonal antibodies. After reversal of the cross-linking, PCR was performed and analyzed as described in panel A to test for the presence of DNA corresponding to the regions of 4/P and 14/T, both of which are modified (thereby were not contained) in the helper plasmid as shown in panels B and G. Left panel (input) shows the results of the PCR conducted on the chromatin before precipitation. (F) The quantification of the raw data shown in panel E. The signals corresponding to each band were quantified by an image analyzer. The ratio of the precipitated signal (IP) to the input signal was calculated for the regions 4/P and 14/T. (G) A schematic diagram of the helper plasmid pM5057. The regions that were amplified by PCR in the ChIP assays (H and I) are indicated as P, T, and I. (H) In vivo binding of HMO1 to the promoter (P), terminator (T) and unrelated intermediate (I) regions of the 35S rRNA gene on the helper plasmid were analyzed by ChIP assays using the same DNA fractions (input and IP) as described in panel E. (I) The quantification of the raw data shown in panel H was performed as described in panel F. WT, wild type.

    Techniques Used: In Vivo, Binding Assay, Chromatin Immunoprecipitation, Expressing, Negative Control, Polymerase Chain Reaction, Polyacrylamide Gel Electrophoresis, Staining, SYBR Green Assay, Plasmid Preparation, Agarose Gel Electrophoresis, Modification, Amplification

    72) Product Images from "Loop-mediated Isothermal Amplification assay for Detection of Candidatus Liberibacter Asiaticus, a Causal Agent of Citrus Huanglongbing"

    Article Title: Loop-mediated Isothermal Amplification assay for Detection of Candidatus Liberibacter Asiaticus, a Causal Agent of Citrus Huanglongbing

    Journal: The Plant Pathology Journal

    doi: 10.5423/PPJ.FT.10.2018.0212

    Sensitivity of loop-mediated isothermal amplification (LAMP), conventional PCR and real-time PCR for detecting HLB. (A) Visual examination of LAMP products by adding SYBR Green I dye. (B) Electrophoresis analysis of LAMP products on 1% agarose gel. (C) Electrophoresis analysis of conventional PCR products on 1% agarose gel. (D) Sensitivity of real-time PCR for detecting HLB using primers (LJ900p, LJ900r)/probe (LJ900p) listed in Table 1 . Tube and lane 1–8, and template DNA for real-time PCR; serially diluted genomic DNA (1, 10 −1 , 10 −2 , 10 −3 , 10 −4 , 10 −5 , 10 −6 ) and distilled water as negative control, respectively, M; 100 bp DNA ladder.
    Figure Legend Snippet: Sensitivity of loop-mediated isothermal amplification (LAMP), conventional PCR and real-time PCR for detecting HLB. (A) Visual examination of LAMP products by adding SYBR Green I dye. (B) Electrophoresis analysis of LAMP products on 1% agarose gel. (C) Electrophoresis analysis of conventional PCR products on 1% agarose gel. (D) Sensitivity of real-time PCR for detecting HLB using primers (LJ900p, LJ900r)/probe (LJ900p) listed in Table 1 . Tube and lane 1–8, and template DNA for real-time PCR; serially diluted genomic DNA (1, 10 −1 , 10 −2 , 10 −3 , 10 −4 , 10 −5 , 10 −6 ) and distilled water as negative control, respectively, M; 100 bp DNA ladder.

    Techniques Used: Amplification, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, SYBR Green Assay, Electrophoresis, Agarose Gel Electrophoresis, Negative Control

    Loop-mediated isothermal amplification (LAMP) for specific detection of ‘ Candidatus Liberibacter asiaticus’ using the primer set from the prophage gene, LasA I in HLB-infected leaves of grapefruit according to reaction temperatures (57, 60, 62 and 65°C). condition test for HLB detection. (A) Visual detection under normal light by adding SYBR Green I dye. (B) Electrophoresis analysis on 1% agarose gel. Lanes 1–4; HLB-infected grapefruit leaves, lanes 5–8; healthy grapefruit leaves, lanes 9–11; distilled water, lane M; 100 bp DNA ladder (NEB New England Biolabs, cat# N3231S).
    Figure Legend Snippet: Loop-mediated isothermal amplification (LAMP) for specific detection of ‘ Candidatus Liberibacter asiaticus’ using the primer set from the prophage gene, LasA I in HLB-infected leaves of grapefruit according to reaction temperatures (57, 60, 62 and 65°C). condition test for HLB detection. (A) Visual detection under normal light by adding SYBR Green I dye. (B) Electrophoresis analysis on 1% agarose gel. Lanes 1–4; HLB-infected grapefruit leaves, lanes 5–8; healthy grapefruit leaves, lanes 9–11; distilled water, lane M; 100 bp DNA ladder (NEB New England Biolabs, cat# N3231S).

    Techniques Used: Amplification, Infection, SYBR Green Assay, Electrophoresis, Agarose Gel Electrophoresis

    73) Product Images from "DNA methyltransferase 3b preferentially associates with condensed chromatin"

    Article Title: DNA methyltransferase 3b preferentially associates with condensed chromatin

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkq870

    Dnmt3b is enriched in histone H1-containing chromatin fractions. ( A ) Dnmt3b, Dnmt3a2, Dnmt3a, Dnmt3L and Oct3/4 expression during ES cell differentiation. Equal amounts of protein (10 µg) from undifferentiated ES cells (Day 0) and differentiated ES cells (from Day 1 to 10) were analyzed by immunoblot using the indicated antibodies. ( B ) Subcellular distribution of Dnmt3b, Dnmt3a2 and Dnmt3L. ES cells were sequentially extracted to obtain cytoplasmic, chromatin and nuclear matrix fractions. Equal amounts of fractionated protein (10 µg) were separated by SDS–PAGE and then analyzed by immunoblot using the indicated antibodies. Fractionation efficiency was confirmed by immunoblot analysis using anti-PCNA (cytoplasm) and anti-lamin B1 (nuclear matrix) antibodies, and by Coomassie Brilliant Blue (CBB) staining of core histones (chromatin). ( C ) Schematic representation of chromatin fractionation. Isolated ES cell nuclei (N) were digested with 5, 20 or 80 U of MNase/mg DNA. Digested nuclei were fractionated into three fractions: transcriptionally active (S1), transcriptionally inactive (S2) and nuclear matrix-containing (P). ( D ) DNA samples from nuclear fractions N, S1, S2 and P. Equivalent amounts of DNA from each fraction were separated by 1.5% agarose gel electrophoresis in 1× TAE followed by SYBR-Green I staining. ( E ) Nuclear aliquots corresponding to 600 ng of DNA from N, S1, S2 and P fractions were separated by SDS–PAGE and then analyzed by immunoblot using the indicated antibodies.
    Figure Legend Snippet: Dnmt3b is enriched in histone H1-containing chromatin fractions. ( A ) Dnmt3b, Dnmt3a2, Dnmt3a, Dnmt3L and Oct3/4 expression during ES cell differentiation. Equal amounts of protein (10 µg) from undifferentiated ES cells (Day 0) and differentiated ES cells (from Day 1 to 10) were analyzed by immunoblot using the indicated antibodies. ( B ) Subcellular distribution of Dnmt3b, Dnmt3a2 and Dnmt3L. ES cells were sequentially extracted to obtain cytoplasmic, chromatin and nuclear matrix fractions. Equal amounts of fractionated protein (10 µg) were separated by SDS–PAGE and then analyzed by immunoblot using the indicated antibodies. Fractionation efficiency was confirmed by immunoblot analysis using anti-PCNA (cytoplasm) and anti-lamin B1 (nuclear matrix) antibodies, and by Coomassie Brilliant Blue (CBB) staining of core histones (chromatin). ( C ) Schematic representation of chromatin fractionation. Isolated ES cell nuclei (N) were digested with 5, 20 or 80 U of MNase/mg DNA. Digested nuclei were fractionated into three fractions: transcriptionally active (S1), transcriptionally inactive (S2) and nuclear matrix-containing (P). ( D ) DNA samples from nuclear fractions N, S1, S2 and P. Equivalent amounts of DNA from each fraction were separated by 1.5% agarose gel electrophoresis in 1× TAE followed by SYBR-Green I staining. ( E ) Nuclear aliquots corresponding to 600 ng of DNA from N, S1, S2 and P fractions were separated by SDS–PAGE and then analyzed by immunoblot using the indicated antibodies.

    Techniques Used: Expressing, Cell Differentiation, SDS Page, Fractionation, Staining, Isolation, Agarose Gel Electrophoresis, SYBR Green Assay

    Dnmt3b associates with nuclease-resistant heterochromatin and the association requires higher-order chromatin structure. ( A ) DNA from nuclei digested with 5 or 80 U MNase/mg DNA. Equivalent amounts of DNA were separated by 1.5% agarose gel electrophoresis in 1× TAE and then visualized with SYBR-green I. ( B ) Detection of chromatin in FLAG-Dnmt3b immunoprecipitates. Nuclear extracts were digested with 5 or 80 U MNase/mg DNA, and subjected to immunoprecipitation using an anti-FLAG antibody in the presence of 75 mM NaCl. FLAG-Dnmt3b immunoprecipitates were analyzed by immunoblot using an anti-histone H3 antibody to confirm the interaction between Dnmt3b and chromatin. Input represents 4% of the nuclear extract used in the assay. ( C ) Dnmt3b co-sedimented with histone H1-containing poly-nucleosomes on a sucrose gradient. S2 fractions were subjected to 10–30% (w/v) sucrose gradient sedimentation. Equivalent aliquots of each DNA fraction and protein sample were analyzed by agarose gel electrophoresis and immunoblot using the indicated antibodies, respectively. Core histones in each fraction were visualized by CBB staining.
    Figure Legend Snippet: Dnmt3b associates with nuclease-resistant heterochromatin and the association requires higher-order chromatin structure. ( A ) DNA from nuclei digested with 5 or 80 U MNase/mg DNA. Equivalent amounts of DNA were separated by 1.5% agarose gel electrophoresis in 1× TAE and then visualized with SYBR-green I. ( B ) Detection of chromatin in FLAG-Dnmt3b immunoprecipitates. Nuclear extracts were digested with 5 or 80 U MNase/mg DNA, and subjected to immunoprecipitation using an anti-FLAG antibody in the presence of 75 mM NaCl. FLAG-Dnmt3b immunoprecipitates were analyzed by immunoblot using an anti-histone H3 antibody to confirm the interaction between Dnmt3b and chromatin. Input represents 4% of the nuclear extract used in the assay. ( C ) Dnmt3b co-sedimented with histone H1-containing poly-nucleosomes on a sucrose gradient. S2 fractions were subjected to 10–30% (w/v) sucrose gradient sedimentation. Equivalent aliquots of each DNA fraction and protein sample were analyzed by agarose gel electrophoresis and immunoblot using the indicated antibodies, respectively. Core histones in each fraction were visualized by CBB staining.

    Techniques Used: Agarose Gel Electrophoresis, SYBR Green Assay, Immunoprecipitation, Sedimentation, Staining

    74) Product Images from "DNA methyltransferase 3b preferentially associates with condensed chromatin"

    Article Title: DNA methyltransferase 3b preferentially associates with condensed chromatin

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkq870

    Dnmt3b is enriched in histone H1-containing chromatin fractions. ( A ) Dnmt3b, Dnmt3a2, Dnmt3a, Dnmt3L and Oct3/4 expression during ES cell differentiation. Equal amounts of protein (10 µg) from undifferentiated ES cells (Day 0) and differentiated ES cells (from Day 1 to 10) were analyzed by immunoblot using the indicated antibodies. ( B ) Subcellular distribution of Dnmt3b, Dnmt3a2 and Dnmt3L. ES cells were sequentially extracted to obtain cytoplasmic, chromatin and nuclear matrix fractions. Equal amounts of fractionated protein (10 µg) were separated by SDS–PAGE and then analyzed by immunoblot using the indicated antibodies. Fractionation efficiency was confirmed by immunoblot analysis using anti-PCNA (cytoplasm) and anti-lamin B1 (nuclear matrix) antibodies, and by Coomassie Brilliant Blue (CBB) staining of core histones (chromatin). ( C ) Schematic representation of chromatin fractionation. Isolated ES cell nuclei (N) were digested with 5, 20 or 80 U of MNase/mg DNA. Digested nuclei were fractionated into three fractions: transcriptionally active (S1), transcriptionally inactive (S2) and nuclear matrix-containing (P). ( D ) DNA samples from nuclear fractions N, S1, S2 and P. Equivalent amounts of DNA from each fraction were separated by 1.5% agarose gel electrophoresis in 1× TAE followed by SYBR-Green I staining. ( E ) Nuclear aliquots corresponding to 600 ng of DNA from N, S1, S2 and P fractions were separated by SDS–PAGE and then analyzed by immunoblot using the indicated antibodies.
    Figure Legend Snippet: Dnmt3b is enriched in histone H1-containing chromatin fractions. ( A ) Dnmt3b, Dnmt3a2, Dnmt3a, Dnmt3L and Oct3/4 expression during ES cell differentiation. Equal amounts of protein (10 µg) from undifferentiated ES cells (Day 0) and differentiated ES cells (from Day 1 to 10) were analyzed by immunoblot using the indicated antibodies. ( B ) Subcellular distribution of Dnmt3b, Dnmt3a2 and Dnmt3L. ES cells were sequentially extracted to obtain cytoplasmic, chromatin and nuclear matrix fractions. Equal amounts of fractionated protein (10 µg) were separated by SDS–PAGE and then analyzed by immunoblot using the indicated antibodies. Fractionation efficiency was confirmed by immunoblot analysis using anti-PCNA (cytoplasm) and anti-lamin B1 (nuclear matrix) antibodies, and by Coomassie Brilliant Blue (CBB) staining of core histones (chromatin). ( C ) Schematic representation of chromatin fractionation. Isolated ES cell nuclei (N) were digested with 5, 20 or 80 U of MNase/mg DNA. Digested nuclei were fractionated into three fractions: transcriptionally active (S1), transcriptionally inactive (S2) and nuclear matrix-containing (P). ( D ) DNA samples from nuclear fractions N, S1, S2 and P. Equivalent amounts of DNA from each fraction were separated by 1.5% agarose gel electrophoresis in 1× TAE followed by SYBR-Green I staining. ( E ) Nuclear aliquots corresponding to 600 ng of DNA from N, S1, S2 and P fractions were separated by SDS–PAGE and then analyzed by immunoblot using the indicated antibodies.

    Techniques Used: Expressing, Cell Differentiation, SDS Page, Fractionation, Staining, Isolation, Agarose Gel Electrophoresis, SYBR Green Assay

    Dnmt3b associates with nuclease-resistant heterochromatin and the association requires higher-order chromatin structure. ( A ) DNA from nuclei digested with 5 or 80 U MNase/mg DNA. Equivalent amounts of DNA were separated by 1.5% agarose gel electrophoresis in 1× TAE and then visualized with SYBR-green I. ( B ) Detection of chromatin in FLAG-Dnmt3b immunoprecipitates. Nuclear extracts were digested with 5 or 80 U MNase/mg DNA, and subjected to immunoprecipitation using an anti-FLAG antibody in the presence of 75 mM NaCl. FLAG-Dnmt3b immunoprecipitates were analyzed by immunoblot using an anti-histone H3 antibody to confirm the interaction between Dnmt3b and chromatin. Input represents 4% of the nuclear extract used in the assay. ( C ) Dnmt3b co-sedimented with histone H1-containing poly-nucleosomes on a sucrose gradient. S2 fractions were subjected to 10–30% (w/v) sucrose gradient sedimentation. Equivalent aliquots of each DNA fraction and protein sample were analyzed by agarose gel electrophoresis and immunoblot using the indicated antibodies, respectively. Core histones in each fraction were visualized by CBB staining.
    Figure Legend Snippet: Dnmt3b associates with nuclease-resistant heterochromatin and the association requires higher-order chromatin structure. ( A ) DNA from nuclei digested with 5 or 80 U MNase/mg DNA. Equivalent amounts of DNA were separated by 1.5% agarose gel electrophoresis in 1× TAE and then visualized with SYBR-green I. ( B ) Detection of chromatin in FLAG-Dnmt3b immunoprecipitates. Nuclear extracts were digested with 5 or 80 U MNase/mg DNA, and subjected to immunoprecipitation using an anti-FLAG antibody in the presence of 75 mM NaCl. FLAG-Dnmt3b immunoprecipitates were analyzed by immunoblot using an anti-histone H3 antibody to confirm the interaction between Dnmt3b and chromatin. Input represents 4% of the nuclear extract used in the assay. ( C ) Dnmt3b co-sedimented with histone H1-containing poly-nucleosomes on a sucrose gradient. S2 fractions were subjected to 10–30% (w/v) sucrose gradient sedimentation. Equivalent aliquots of each DNA fraction and protein sample were analyzed by agarose gel electrophoresis and immunoblot using the indicated antibodies, respectively. Core histones in each fraction were visualized by CBB staining.

    Techniques Used: Agarose Gel Electrophoresis, SYBR Green Assay, Immunoprecipitation, Sedimentation, Staining

    75) Product Images from "Frequent fusion and fission of plant mitochondria with unequal nucleoid distribution"

    Article Title: Frequent fusion and fission of plant mitochondria with unequal nucleoid distribution

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    doi: 10.1073/pnas.0401077101

    Mitochondria and mitochondrial nucleoids in tobacco suspension cultured cells (BY-2). ( a ) Mitochondria stained by MitoTracker ( Left ) and mitochondrial nucleoids stained by Sybr Green I ( Right ). In the merged images ( Center ), the nucleoids appear yellow because of the combination of red and green fluorescence. Arrowheads indicate mitochondria that have no visible nucleoids. ( b ) Views of three mitochondria at 0.5-μm intervals along the optical axis. Yellow nucleoids are visible in the lower two mitochondria but not in the top mitochondria. ( c and d ) Time series of fission of mitochondria with nucleoids shown at 10-sec intervals. (Scale bars, 2 μm.) Movies 2–4.
    Figure Legend Snippet: Mitochondria and mitochondrial nucleoids in tobacco suspension cultured cells (BY-2). ( a ) Mitochondria stained by MitoTracker ( Left ) and mitochondrial nucleoids stained by Sybr Green I ( Right ). In the merged images ( Center ), the nucleoids appear yellow because of the combination of red and green fluorescence. Arrowheads indicate mitochondria that have no visible nucleoids. ( b ) Views of three mitochondria at 0.5-μm intervals along the optical axis. Yellow nucleoids are visible in the lower two mitochondria but not in the top mitochondria. ( c and d ) Time series of fission of mitochondria with nucleoids shown at 10-sec intervals. (Scale bars, 2 μm.) Movies 2–4.

    Techniques Used: Cell Culture, Staining, SYBR Green Assay, Fluorescence, Size-exclusion Chromatography

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    Transformation Assay:

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    Transferring:

    Article Title: Transcriptome analysis of functional differentiation between haploid and diploid cells of Emiliania huxleyi, a globally significant photosynthetic calcifying cell
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    Imaging:

    Article Title: Inducing controlled cell cycle arrest and re-entry during asexual proliferation of Plasmodium falciparum malaria parasites
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    Polymerase Chain Reaction:

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    In Vivo:

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    Article Snippet: Paragraph title: Analysis of in vivo -bound gp16-DNA complex ... The gel was stained with SYBR Green I (Life Technologies, Thermo Fisher Scientific, Inc.) for DNA and then with Coomassie blue R-250 for protein.

    Fluorescence:

    Article Title: Evaluation of a Direct Reverse Transcription Loop-Mediated Isothermal Amplification Method without RNA Extraction for the Detection of Human Enterovirus 71 Subgenotype C4 in Nasopharyngeal Swab Specimens
    Article Snippet: Direct RT-LAMP was performed the same as RT-LAMP described above except the incubation time was extended to 75 min and 1 μl of 1∶100 diluted SYBR green I (Invitrogen, Eugene Oregon, USA) was added after amplification for the observation under the UV light by naked eyes. .. Positive reactions were defined as those samples having a threshold value of greater than 0.2 or a color change from orange to green fluorescence.

    RT Lamp Assay:

    Article Title: Evaluation of a Direct Reverse Transcription Loop-Mediated Isothermal Amplification Method without RNA Extraction for the Detection of Human Enterovirus 71 Subgenotype C4 in Nasopharyngeal Swab Specimens
    Article Snippet: .. Similarly, the specificity and sensitivity of both the RT-LAMP and the direct RT-LAMP were performed using the Loopamp RNA amplification kit and the fluorescent detection reagent (FDR) (Eiken Chemical Co., Ltd., Tokyo, Japan) in accordance with the manufacturer's instructions with the exception of the addition of a 1∶100 diluted SYBR Green I (Invitrogen, Eugene Oregon, USA) in place of the FDR after the amplification for the direct RT-LAMP assay. ..

    Isolation:

    Article Title: Transcriptome analysis of functional differentiation between haploid and diploid cells of Emiliania huxleyi, a globally significant photosynthetic calcifying cell
    Article Snippet: A clonal 1N strain (RCC1217) was isolated from RCC1216 following a partial phase change (2N to 1N; Table ). .. Absence of bacteria was confirmed by inoculation in bacterial test media (protocols available at the Center for Culture of Marine Phytoplankton [ ]) and by extensive epifluorescence examination of samples fixed in 1% formaldehyde, 0.05% glutaraldehyde, stained with Sybr Green I (Invitrogen, Carlsbad, California, USA), and collected onto 0.2 μm pore-size filters (Millipore).

    Size-exclusion Chromatography:

    Article Title: A high-throughput qPCR system for simultaneous quantitative detection of dairy Lactococcus lactis and Leuconostoc bacteriophages
    Article Snippet: To test the optimum concentrations of the primers, identical reactions containing different concentrations of the forward and reverse primers, prepared essentially as described [ ], were run using the following thermocycling conditions: initial stage at 50°C for 2 min, hot start at 95°C for 2 min; followed by 40 cycles of (i) 95°C for 15 sec, (ii) 55°C for 30 sec and (iii) 72°C for 30 sec. .. PCR reactions for comparison of SYBR Green I (Thermo Fisher Scientific) and EvaGreen (Biotium, USA) detection chemistries were prepared as described above using the P220 phage DNA as template.

    Microscopy:

    Article Title: Inducing controlled cell cycle arrest and re-entry during asexual proliferation of Plasmodium falciparum malaria parasites
    Article Snippet: The effect of DFMO on nuclear division was confirmed by fluorescent microscopy by removing samples from treated and control parasites as above before methanol fixation on microscope slides and staining with 10 µl 1:1000 SYBR Green I for 30 min in the dark at room temperature. .. Samples were stained with 10 µl 1:1000 SYBR Green I (Invitrogen) for DNA quantification and subsequently with 10 μl Pyronin Y (100 μg/ml) for RNA quantification for 30 min each in the dark at room temperature, as has been described .

    Purification:

    Article Title: Exclusion of small terminase mediated DNA threading models for genome packaging in bacteriophage T4
    Article Snippet: The gp16 proteins purified as above were electrophoresed on a 4–20% gradient native polyacrylamide gel (Life Technologies, Thermo Fisher Scientific, Inc.) under non-denaturing conditions using 1× Tris/borate/EDTA buffer (89 mM Tris–HCl, pH 7.6, 89 mM boric acid, and 2 mM EDTA). .. The gel was stained with SYBR Green I (Life Technologies, Thermo Fisher Scientific, Inc.) for DNA and then with Coomassie blue R-250 for protein.

    Labeling:

    Article Title: An Adaptable Two-Color Flow Cytometric Assay to Quantitate the Invasion of Erythrocytes by Plasmodium falciparum Parasites
    Article Snippet: Paragraph title: Parasite Labeling ... The cells were washed with PBS before staining with 10 μg/mL ethidium bromide (Sigma-Aldrich, Dorset, UK) in PBS, 2 μM Hoechst 33342 (Invitrogen, Paisley, UK) in RPMI 1640 or 1:5,000 SYBR® Green I (Invitrogen, Paisley, UK) in PBS, for 1 h at 37°C.

    Article Title: Targeted Sorting of Single Virus-Infected Cells of the Coccolithophore Emiliania huxleyi
    Article Snippet: .. 1.4×105 cells ml∼1 ) were placed into microcentrifuge Eppendorf tubes and labeled with either fluorescent dye N-(3-riethylammoniumpropyl)-4-[4-(dibutylamino)styryl] pyridinium dibromide (FM 1-43, Invitrogen Co., Carlsbad, CA, USA), 5-(and-6)-chloromethyl-2′,7′-dichlorodihydrofluorescein diacetate (CM-H2 DCFDA, Molecular Probes Inc., Eugene, OR, USA) or SYBR Green I (Molecular Probes Inc., Eugene, OR, USA). .. The cells were incubated with 10 µM (final concentration) FM 1-43 , 5 mM (final concentration) CM-H2 DCFDA or SYBR Green I (5×105 dilution of commercial stock) for 30 min, 60 min or 15 min, respectively, in the dark at 15°C.

    Article Title: Monitoring of Dynamic Microbiological Processes Using Real-Time Flow Cytometry
    Article Snippet: Paragraph title: Fluorescent labeling of heat-induced cell damage ... A 5 mL water sample was pre-warmed (35 °C, 10 minutes) and then stained with a 10 µL/mL of a stock solution of SYBR Green I (SG; Invitrogen) and propidium iodide (PI; Invitrogen), comprising 100x diluted SG (from original stock solution of 10’000x concentrate) and 300 µM PI in TRIS Buffer [ ].

    FACS:

    Article Title: Inducing controlled cell cycle arrest and re-entry during asexual proliferation of Plasmodium falciparum malaria parasites
    Article Snippet: Analyses were performed on a Becton Dickenson FACS ARIA with 515–545 nm band pass filters (FL-1 channel, FITC signal) for SYBR Green I (DNA) and analysed using FlowJo vX.0.7, with gating as described in the Supplementary Methods. .. Samples were stained with 10 µl 1:1000 SYBR Green I (Invitrogen) for DNA quantification and subsequently with 10 μl Pyronin Y (100 μg/ml) for RNA quantification for 30 min each in the dark at room temperature, as has been described .

    Plasmid Preparation:

    Article Title: Frequent fusion and fission of plant mitochondria with unequal nucleoid distribution
    Article Snippet: Tobacco BY-2 cells were chosen because they stain well with both MitoTracker (Molecular Probes) and SYBR Green I (Molecular Probes). .. Plasmid DNA was transiently introduced into tobacco BY-2 cells and onion epidermal cells on agar plates by a helium-driven particle accelerator (PDS/1000; Bio-Rad) with all basic adjustments set according to the manufacturer's recommendations.

    Electrophoresis:

    Article Title: Application of a loop-mediated isothermal amplification (LAMP) assay targeting cox1 gene for the detection of Clonorchis sinensis in human fecal samples
    Article Snippet: Amplified LAMP products were detected by adding 1.0 μl of 1:10 diluted 10,000x concentration of SYBR Green I (Invitrogen, Carlsbad, CA, USA) to each tube. .. In addition, 5.0 μl of the LAMP products was examined by electrophoresis on a 2% agarose gel, followed by ethidium bromide staining and visualization under UV light.

    RNA Extraction:

    Article Title: Evaluation of a Direct Reverse Transcription Loop-Mediated Isothermal Amplification Method without RNA Extraction for the Detection of Human Enterovirus 71 Subgenotype C4 in Nasopharyngeal Swab Specimens
    Article Snippet: Paragraph title: Direct RT-LAMP with heat-treated samples without RNA extraction ... Direct RT-LAMP was performed the same as RT-LAMP described above except the incubation time was extended to 75 min and 1 μl of 1∶100 diluted SYBR green I (Invitrogen, Eugene Oregon, USA) was added after amplification for the observation under the UV light by naked eyes.

    Agarose Gel Electrophoresis:

    Article Title: Application of a loop-mediated isothermal amplification (LAMP) assay targeting cox1 gene for the detection of Clonorchis sinensis in human fecal samples
    Article Snippet: Amplified LAMP products were detected by adding 1.0 μl of 1:10 diluted 10,000x concentration of SYBR Green I (Invitrogen, Carlsbad, CA, USA) to each tube. .. In addition, 5.0 μl of the LAMP products was examined by electrophoresis on a 2% agarose gel, followed by ethidium bromide staining and visualization under UV light.

    Article Title: Rapid Detection of Zika Virus in Urine Samples and Infected Mosquitos by Reverse Transcription-Loop-Mediated Isothermal Amplification
    Article Snippet: One half of the RT-LAMP reaction was electrophoresed along with Invitrogen Low DNA Mass Ladder on a 1 × Nancy-520, 2% agarose gel in 1 × TAE buffer (40 mM Tris, 20 mM acetic acid, 1 mM EDTA) at 90 V for 90–120 minutes. .. In addition, a 1:10 SYBR green I (Life Technologies) dilution was made in TAE buffer, then 2.0 μL of the SYBR dilution was added to the remaining half of the reaction.

    Laser-Scanning Microscopy:

    Article Title: Inducing controlled cell cycle arrest and re-entry during asexual proliferation of Plasmodium falciparum malaria parasites
    Article Snippet: Stained samples were visualised using Zeiss LSM 880 Confocal Laser Scanning Microscope (LSM) - Airyscan detector for super-resolution imaging using a 488 nm laser and 495–550 BP filter. .. Samples were stained with 10 µl 1:1000 SYBR Green I (Invitrogen) for DNA quantification and subsequently with 10 μl Pyronin Y (100 μg/ml) for RNA quantification for 30 min each in the dark at room temperature, as has been described .

    Sampling:

    Article Title: Monitoring of Dynamic Microbiological Processes Using Real-Time Flow Cytometry
    Article Snippet: A 5 mL water sample was pre-warmed (35 °C, 10 minutes) and then stained with a 10 µL/mL of a stock solution of SYBR Green I (SG; Invitrogen) and propidium iodide (PI; Invitrogen), comprising 100x diluted SG (from original stock solution of 10’000x concentrate) and 300 µM PI in TRIS Buffer [ ]. .. The sample vial was connected directly to the FCM via the instrument’s sampling port.

    Concentration Assay:

    Article Title: One-Pot Reverse Transcriptional Loop-Mediated Isothermal Amplification (RT-LAMP) for Detecting MERS-CoV
    Article Snippet: .. One-Pot RT-LAMP in Microchamber In order to utilize EvaGreen in one-pot RT-LAMP experiment, the sensitivity of EvaGreen® (Biotium Inc., USA) were compared with SYBR® Green I (Thermo Fisher Scientific, Waltham, MA, USA) for each concentration (non-contained, 1, 2, 5, 10, 20, 50, 100, 200, 500 and 1000X). .. Polymer based microchamber devices were purchased from Dongwoo Science Co., Ltd (Republic of Korea).

    Article Title: Application of a loop-mediated isothermal amplification (LAMP) assay targeting cox1 gene for the detection of Clonorchis sinensis in human fecal samples
    Article Snippet: .. Amplified LAMP products were detected by adding 1.0 μl of 1:10 diluted 10,000x concentration of SYBR Green I (Invitrogen, Carlsbad, CA, USA) to each tube. .. The amplicon was observed directly either by the naked eye or by placing the reaction tube under UV light (Gel documentation system, UVItech, Cambridge, UK).

    Article Title: Targeted Sorting of Single Virus-Infected Cells of the Coccolithophore Emiliania huxleyi
    Article Snippet: Fluorescent cell labeling 1 ml E. huxleyi culture aliquots (concentration was adjusted to approx. .. 1.4×105 cells ml∼1 ) were placed into microcentrifuge Eppendorf tubes and labeled with either fluorescent dye N-(3-riethylammoniumpropyl)-4-[4-(dibutylamino)styryl] pyridinium dibromide (FM 1-43, Invitrogen Co., Carlsbad, CA, USA), 5-(and-6)-chloromethyl-2′,7′-dichlorodihydrofluorescein diacetate (CM-H2 DCFDA, Molecular Probes Inc., Eugene, OR, USA) or SYBR Green I (Molecular Probes Inc., Eugene, OR, USA).

    Lamp Assay:

    Article Title: Application of a loop-mediated isothermal amplification (LAMP) assay targeting cox1 gene for the detection of Clonorchis sinensis in human fecal samples
    Article Snippet: The LAMP reaction The LAMP assay was performed in a total reaction mixture volume of 25 μl, containing 2.5 μl of 10x ThermoPol reaction buffer (New England BioLabs, Ipswich, MA, USA), 6 mM MgSO4 , 1 M betaine, 1.4 mM dNTP mix, 0.2 μM each of F3 and B3 primers, 1.6 μM each of FIP and BIP primers, 0.8 μM each of LF and LB primers, 8 U Bst DNA polymerase (New England BioLabs) and 4 μl of the template DNA. .. Amplified LAMP products were detected by adding 1.0 μl of 1:10 diluted 10,000x concentration of SYBR Green I (Invitrogen, Carlsbad, CA, USA) to each tube.

    Staining:

    Article Title: An Adaptable Two-Color Flow Cytometric Assay to Quantitate the Invasion of Erythrocytes by Plasmodium falciparum Parasites
    Article Snippet: .. The cells were washed with PBS before staining with 10 μg/mL ethidium bromide (Sigma-Aldrich, Dorset, UK) in PBS, 2 μM Hoechst 33342 (Invitrogen, Paisley, UK) in RPMI 1640 or 1:5,000 SYBR® Green I (Invitrogen, Paisley, UK) in PBS, for 1 h at 37°C. .. After staining, the cells were washed with PBS, before being fixed with a 2% paraformaldehyde (Sigma-Aldrich, Dorset, UK), 0.2% glutaraldehyde (Sigma-Aldrich, Dorset, UK) solution in PBS for 1 h at 4°C.

    Article Title: Effectiveness of Stevia Rebaudiana Whole Leaf Extract Against the Various Morphological Forms of Borrelia Burgdorferi in Vitro
    Article Snippet: To 1 ml of sterilized distilled water, 10 μl of SYBR Green I (10,000× stock, Invitrogen, Grand Island, NY) and 30 μl of propidium iodide (Thermo Scientific) were briefly mixed. .. The staining mixture (10 μl) was added to all the wells containing B. burgdorferi and was incubated in the dark for 15 min.

    Article Title: Exclusion of small terminase mediated DNA threading models for genome packaging in bacteriophage T4
    Article Snippet: .. The gel was stained with SYBR Green I (Life Technologies, Thermo Fisher Scientific, Inc.) for DNA and then with Coomassie blue R-250 for protein. .. Analysis of gp16 binding to DNA in vitro The gp16 proteins (20–75 μM) purified as above were mixed with the 500-bp PCR amplified g16 DNA (5 nM) in a buffer containing 15 mM Tris–HCl, pH 8 and 75 mM NaCl) and incubated at room temperature for 15 min.

    Article Title: Application of a loop-mediated isothermal amplification (LAMP) assay targeting cox1 gene for the detection of Clonorchis sinensis in human fecal samples
    Article Snippet: Amplified LAMP products were detected by adding 1.0 μl of 1:10 diluted 10,000x concentration of SYBR Green I (Invitrogen, Carlsbad, CA, USA) to each tube. .. In addition, 5.0 μl of the LAMP products was examined by electrophoresis on a 2% agarose gel, followed by ethidium bromide staining and visualization under UV light.

    Article Title: Inducing controlled cell cycle arrest and re-entry during asexual proliferation of Plasmodium falciparum malaria parasites
    Article Snippet: .. Samples were stained with 10 µl 1:1000 SYBR Green I (Invitrogen) for DNA quantification and subsequently with 10 μl Pyronin Y (100 μg/ml) for RNA quantification for 30 min each in the dark at room temperature, as has been described . .. Analyses were performed for SYBR Green I (DNA) as described above with Pyronin Y (RNA) measured at 564–606 nm (FL-2 channel, PE signal, 10 000 events of parasitised cells captured).

    Article Title: Monitoring of Dynamic Microbiological Processes Using Real-Time Flow Cytometry
    Article Snippet: .. A 5 mL water sample was pre-warmed (35 °C, 10 minutes) and then stained with a 10 µL/mL of a stock solution of SYBR Green I (SG; Invitrogen) and propidium iodide (PI; Invitrogen), comprising 100x diluted SG (from original stock solution of 10’000x concentrate) and 300 µM PI in TRIS Buffer [ ]. .. The stained sample was incubated (35 °C, 10 minutes) prior to the start of the RT-FCM measurements to allow complete staining of all bacterial cells.

    Article Title: Frequent fusion and fission of plant mitochondria with unequal nucleoid distribution
    Article Snippet: .. Tobacco BY-2 cells were chosen because they stain well with both MitoTracker (Molecular Probes) and SYBR Green I (Molecular Probes). .. Plasmid DNA was transiently introduced into tobacco BY-2 cells and onion epidermal cells on agar plates by a helium-driven particle accelerator (PDS/1000; Bio-Rad) with all basic adjustments set according to the manufacturer's recommendations.

    Article Title: Transcriptome analysis of functional differentiation between haploid and diploid cells of Emiliania huxleyi, a globally significant photosynthetic calcifying cell
    Article Snippet: .. Absence of bacteria was confirmed by inoculation in bacterial test media (protocols available at the Center for Culture of Marine Phytoplankton [ ]) and by extensive epifluorescence examination of samples fixed in 1% formaldehyde, 0.05% glutaraldehyde, stained with Sybr Green I (Invitrogen, Carlsbad, California, USA), and collected onto 0.2 μm pore-size filters (Millipore). ..

    Article Title: Counting Viruses and Bacteria in Photosynthetic Microbial Mats
    Article Snippet: After staining, the filters were placed in glass slides with an antifade solution consisting of 50%:50% (vol/vol) glycerol-PBS (0.05 M Na2 HPO4 , 0.85% NaCl [pH 7.5]) with 1% p -phenylenediamine (Sigma-Aldrich, The Netherlands). .. Two different nucleic acid-specific fluorescent dyes, SYBR gold and SYBR green I ( ) (Life Technologies, NY), were tested.

    Epifluorescence Microscopy:

    Article Title: Counting Viruses and Bacteria in Photosynthetic Microbial Mats
    Article Snippet: Paragraph title: Epifluorescence microscopy. ... Two different nucleic acid-specific fluorescent dyes, SYBR gold and SYBR green I ( ) (Life Technologies, NY), were tested.

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    Thermo Fisher sybr green i
    (A) The RT-LAMP detection results based on different ratios of Bst polymerase and M-MLV reverse transcriptase concentration (100 unit). Concentration of Bst polymerse is 1, 2, 4, 6, and 8 units. (B) Effect of reaction time for RT-LAMP reaction. DNA ladder-like pattern was confirmed by 2% agarose gel electrophoresis. LAMP products detected by adding 1,000X <t>SYBR-green</t> I when the reaction was completed (UV/on and off). Land M: 100 bp DNA ladders; - depicts tube used as negative control, non-template; + indicates the results with template.
    Sybr Green I, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 803 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher sybr green i fluorescence emission
    Microwave-assisted rolling circle amplification reaction mixtures containing 4-fold increased concentrations of one component. (a) Agarose gel electrophoresis results. (b) Fluorescence intensity of <t>SYBR</t> Green I. C = conventional, MW = microwave.
    Sybr Green I Fluorescence Emission, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 77/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    (A) The RT-LAMP detection results based on different ratios of Bst polymerase and M-MLV reverse transcriptase concentration (100 unit). Concentration of Bst polymerse is 1, 2, 4, 6, and 8 units. (B) Effect of reaction time for RT-LAMP reaction. DNA ladder-like pattern was confirmed by 2% agarose gel electrophoresis. LAMP products detected by adding 1,000X SYBR-green I when the reaction was completed (UV/on and off). Land M: 100 bp DNA ladders; - depicts tube used as negative control, non-template; + indicates the results with template.

    Journal: Frontiers in Microbiology

    Article Title: One-Pot Reverse Transcriptional Loop-Mediated Isothermal Amplification (RT-LAMP) for Detecting MERS-CoV

    doi: 10.3389/fmicb.2016.02166

    Figure Lengend Snippet: (A) The RT-LAMP detection results based on different ratios of Bst polymerase and M-MLV reverse transcriptase concentration (100 unit). Concentration of Bst polymerse is 1, 2, 4, 6, and 8 units. (B) Effect of reaction time for RT-LAMP reaction. DNA ladder-like pattern was confirmed by 2% agarose gel electrophoresis. LAMP products detected by adding 1,000X SYBR-green I when the reaction was completed (UV/on and off). Land M: 100 bp DNA ladders; - depicts tube used as negative control, non-template; + indicates the results with template.

    Article Snippet: After addition of SYBR-green I, samples that turned yellowish green were considered positive.

    Techniques: Concentration Assay, Agarose Gel Electrophoresis, SYBR Green Assay, Negative Control

    Microwave-assisted rolling circle amplification reaction mixtures containing 4-fold increased concentrations of one component. (a) Agarose gel electrophoresis results. (b) Fluorescence intensity of SYBR Green I. C = conventional, MW = microwave.

    Journal: PLoS ONE

    Article Title: Controlled Microwave Heating Accelerates Rolling Circle Amplification

    doi: 10.1371/journal.pone.0136532

    Figure Lengend Snippet: Microwave-assisted rolling circle amplification reaction mixtures containing 4-fold increased concentrations of one component. (a) Agarose gel electrophoresis results. (b) Fluorescence intensity of SYBR Green I. C = conventional, MW = microwave.

    Article Snippet: Fluorescence measurements The intensity of SYBR Green I fluorescence emission was measured using a fluorescence plate reader (FluoroCount, Packard) with a 96-well white microwell plate (Thermo Scientific).

    Techniques: Amplification, Agarose Gel Electrophoresis, Fluorescence, SYBR Green Assay

    DFMO arrest and putrescine reversal associated to G1/S cell cycle control in intraerythrocytic P. falciparum parasites. In all instances, ■ refers to control parasites, ▼ refers to DFMO-treated parasites, and ● refers to putrescine-reversed parasites. ( a ) DFMO dose-response curves of asexual P . falciparum 3D7 proliferation over 96 h (initiated with ring-stage parasites, 1% haematocrit, 1% parasitaemia) at 37 °C in the absence or presence of 2 mM putrescine. Proliferation is expressed relative to untreated controls, with data averaged from n = 6 biological replicates and shown ± S.E. ( b ) Synchronised P . falciparum 3D7 cultures were treated with DFMO alone (IC 90 ) or with putrescine (2 mM, after 24 h DFMO pressure) and parasitaemia monitored over 96 h with SYBR Green I fluorescence (10 000 infected erythrocytes counted). * P

    Journal: Scientific Reports

    Article Title: Inducing controlled cell cycle arrest and re-entry during asexual proliferation of Plasmodium falciparum malaria parasites

    doi: 10.1038/s41598-018-34964-w

    Figure Lengend Snippet: DFMO arrest and putrescine reversal associated to G1/S cell cycle control in intraerythrocytic P. falciparum parasites. In all instances, ■ refers to control parasites, ▼ refers to DFMO-treated parasites, and ● refers to putrescine-reversed parasites. ( a ) DFMO dose-response curves of asexual P . falciparum 3D7 proliferation over 96 h (initiated with ring-stage parasites, 1% haematocrit, 1% parasitaemia) at 37 °C in the absence or presence of 2 mM putrescine. Proliferation is expressed relative to untreated controls, with data averaged from n = 6 biological replicates and shown ± S.E. ( b ) Synchronised P . falciparum 3D7 cultures were treated with DFMO alone (IC 90 ) or with putrescine (2 mM, after 24 h DFMO pressure) and parasitaemia monitored over 96 h with SYBR Green I fluorescence (10 000 infected erythrocytes counted). * P

    Article Snippet: DFMO dose-response analyses in the presence and absence of putrescine dihydrochloride (2 mM) were initiated with ring-stage intraerythrocytic P . falciparum parasite cultures (1% haematocrit, 1% parasitaemia) using SYBR Green I fluorescence measured with a Fluoroskan Ascent FL microplate fluorometer (Thermo Scientific, excitation at 485 nm and emission at 538 nm).

    Techniques: SYBR Green Assay, Fluorescence, Infection