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    Name:
    Qubit dsDNA HS Assay Kit
    Description:
    The Qubit dsDNA HS High Sensitivity Assay Kit is designed specifically for use with the Qubit Fluorometer The assay is highly selective for double stranded DNA dsDNA over RNA and is designed to be accurate for initial sample concentrations from 10 pg µL to 100 ng µL The kit provides concentrated assay reagent dilution buffer and pre diluted DNA standards Simply dilute the reagent using the buffer provided add your sample any volume between 1 µl and 20 µl is acceptable and read the concentration using the Qubit Fluorometer Common contaminants such as salts free nucleotides solvents detergents or protein are well tolerated in the assay Which product to choose for dsDNA HS High Sensitivity quantitation • For 1 20 samples use this Qubit dsDNA HS Assay Kit with the Qubit Fluorometer • For 20 2000 samples use the Quant iT dsDNA HS Assay Kit with microplate readerNotes 1 All Qubit assay kits can be used with the Qubit 1 0 Qubit 2 0 Qubit 3 and Qubit 4 fluorometers 2 500 µL thin walled PCR tubes are required but not included 3 Also available as a 1X dsDNA assay kit that provides reagent and buffer in a formulation that is stable as a ready to use solution for up to one year after preparation
    Catalog Number:
    q32851
    Price:
    None
    Applications:
    DNA & RNA Purification & Analysis|DNA Quantitation|Nucleic Acid Quantitation
    Category:
    Kits and Assays
    Buy from Supplier


    Structured Review

    Thermo Fisher rna
    Ureaplasma -driven pyroptosis in HBMEC. Key genes in pyroptosis (Fig. 1 ) were assessed for mRNA responses upon stimulation of HBMEC for 4 h and 30 h. Caspase 1 mRNA expression was determined via <t>RNA</t> sequencing ( a ) and <t>qRT-PCR</t> ( b ). Similarly, RNA sequencing ( c ) and qRT-PCR ( d ) were used to assess caspase 4 mRNA levels. RNA sequencing furthermore determined mRNA expression of NLRP3 ( e ) and gasdermin D ( f ). Data are presented as means ± SD (* p
    The Qubit dsDNA HS High Sensitivity Assay Kit is designed specifically for use with the Qubit Fluorometer The assay is highly selective for double stranded DNA dsDNA over RNA and is designed to be accurate for initial sample concentrations from 10 pg µL to 100 ng µL The kit provides concentrated assay reagent dilution buffer and pre diluted DNA standards Simply dilute the reagent using the buffer provided add your sample any volume between 1 µl and 20 µl is acceptable and read the concentration using the Qubit Fluorometer Common contaminants such as salts free nucleotides solvents detergents or protein are well tolerated in the assay Which product to choose for dsDNA HS High Sensitivity quantitation • For 1 20 samples use this Qubit dsDNA HS Assay Kit with the Qubit Fluorometer • For 20 2000 samples use the Quant iT dsDNA HS Assay Kit with microplate readerNotes 1 All Qubit assay kits can be used with the Qubit 1 0 Qubit 2 0 Qubit 3 and Qubit 4 fluorometers 2 500 µL thin walled PCR tubes are required but not included 3 Also available as a 1X dsDNA assay kit that provides reagent and buffer in a formulation that is stable as a ready to use solution for up to one year after preparation
    https://www.bioz.com/result/rna/product/Thermo Fisher
    Average 94 stars, based on 71218 article reviews
    Price from $9.99 to $1999.99
    rna - by Bioz Stars, 2020-08
    94/100 stars

    Images

    1) Product Images from "More than just inflammation: Ureaplasma species induce apoptosis in human brain microvascular endothelial cells"

    Article Title: More than just inflammation: Ureaplasma species induce apoptosis in human brain microvascular endothelial cells

    Journal: Journal of Neuroinflammation

    doi: 10.1186/s12974-019-1413-8

    Ureaplasma -driven pyroptosis in HBMEC. Key genes in pyroptosis (Fig. 1 ) were assessed for mRNA responses upon stimulation of HBMEC for 4 h and 30 h. Caspase 1 mRNA expression was determined via RNA sequencing ( a ) and qRT-PCR ( b ). Similarly, RNA sequencing ( c ) and qRT-PCR ( d ) were used to assess caspase 4 mRNA levels. RNA sequencing furthermore determined mRNA expression of NLRP3 ( e ) and gasdermin D ( f ). Data are presented as means ± SD (* p
    Figure Legend Snippet: Ureaplasma -driven pyroptosis in HBMEC. Key genes in pyroptosis (Fig. 1 ) were assessed for mRNA responses upon stimulation of HBMEC for 4 h and 30 h. Caspase 1 mRNA expression was determined via RNA sequencing ( a ) and qRT-PCR ( b ). Similarly, RNA sequencing ( c ) and qRT-PCR ( d ) were used to assess caspase 4 mRNA levels. RNA sequencing furthermore determined mRNA expression of NLRP3 ( e ) and gasdermin D ( f ). Data are presented as means ± SD (* p

    Techniques Used: Expressing, RNA Sequencing Assay, Quantitative RT-PCR

    Ureaplasma -driven apoptosis in HBMEC. Enzymes and other proteins involved in the apoptotic cascade (Fig. 1 ) were analyzed upon stimulation of HBMEC for 4 h, 24 h, or 30 h. For caspase 3, mRNA expression was determined via RNA sequencing ( a ) and qRT-PCR ( b ) and enzyme activity (cleaved caspase 3) was assessed via flow cytometry ( c ). For caspase 8, RNA sequencing ( d ) and qRT-PCR ( e ) were used to evaluate mRNA levels and flow cytometry ( f ) was employed to determine protein expression. Caspase 9 mRNA levels were also assessed via RNA sequencing ( g ) and qRT-PCR ( h ), and levels of active caspase 9 were determined using flow cytometry ( i ). RNA sequencing was used to assess mRNA expression of caspase 7 ( j ), BAK ( k ), BAX ( l ), p53 ( m ), FOS ( n ), and APAF1 ( o ). Data are shown as means ± SD (* p
    Figure Legend Snippet: Ureaplasma -driven apoptosis in HBMEC. Enzymes and other proteins involved in the apoptotic cascade (Fig. 1 ) were analyzed upon stimulation of HBMEC for 4 h, 24 h, or 30 h. For caspase 3, mRNA expression was determined via RNA sequencing ( a ) and qRT-PCR ( b ) and enzyme activity (cleaved caspase 3) was assessed via flow cytometry ( c ). For caspase 8, RNA sequencing ( d ) and qRT-PCR ( e ) were used to evaluate mRNA levels and flow cytometry ( f ) was employed to determine protein expression. Caspase 9 mRNA levels were also assessed via RNA sequencing ( g ) and qRT-PCR ( h ), and levels of active caspase 9 were determined using flow cytometry ( i ). RNA sequencing was used to assess mRNA expression of caspase 7 ( j ), BAK ( k ), BAX ( l ), p53 ( m ), FOS ( n ), and APAF1 ( o ). Data are shown as means ± SD (* p

    Techniques Used: Expressing, RNA Sequencing Assay, Quantitative RT-PCR, Activity Assay, Flow Cytometry, Cytometry

    2) Product Images from "Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application"

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application

    Journal: BMC Genomics

    doi: 10.1186/s12864-017-4371-5

    Storage condition of purified ChIP DNA is important. Purified ChIP DNA was adjusted to a concentration of 1 ng/μL ( a ) or 0.1 ng/μL ( b ), aliquoted into 4 different types of microcentrifuge tubes in 15 μL volume, and stored at −20 °C. DNA was quantified using Qubit dsDNA High Sensitivity assay at the indicated time points and expressed as a percentage of the amount measured at day 0. Three independent DNA samples were used in the experiment and DNA concentration from five tubes were measured at each time point. MaxyClear, Axygen® 1.7 mL MaxyClear Snaplock Microcentrifuge Tube; LoBind, Eppendorf DNA LoBind Snap Cap PCR Tube; Siliconized, Fisherbrand™ Siliconized Low-Retention Microcentrifuge Tube; Premium, Fisherbrand™ Premium Microcentrifuge Tube
    Figure Legend Snippet: Storage condition of purified ChIP DNA is important. Purified ChIP DNA was adjusted to a concentration of 1 ng/μL ( a ) or 0.1 ng/μL ( b ), aliquoted into 4 different types of microcentrifuge tubes in 15 μL volume, and stored at −20 °C. DNA was quantified using Qubit dsDNA High Sensitivity assay at the indicated time points and expressed as a percentage of the amount measured at day 0. Three independent DNA samples were used in the experiment and DNA concentration from five tubes were measured at each time point. MaxyClear, Axygen® 1.7 mL MaxyClear Snaplock Microcentrifuge Tube; LoBind, Eppendorf DNA LoBind Snap Cap PCR Tube; Siliconized, Fisherbrand™ Siliconized Low-Retention Microcentrifuge Tube; Premium, Fisherbrand™ Premium Microcentrifuge Tube

    Techniques Used: Purification, Chromatin Immunoprecipitation, Concentration Assay, Sensitive Assay, Polymerase Chain Reaction

    3) Product Images from "Macrophage Coordination of the Interferon Lambda Immune Response"

    Article Title: Macrophage Coordination of the Interferon Lambda Immune Response

    Journal: Frontiers in Immunology

    doi: 10.3389/fimmu.2019.02674

    Monocyte differentiation with IFN-λ3 drives a pro-inflammatory macrophage phenotype. Monocytes were cultured with M- or GM-CSF ± IFN-λ3 for 7 days to examine the effect of IFN-λ3 on Mϕ differentiation, followed by phenotypic and functional characterization (A) . RNA sequencing ( n = 3/treatment) demonstrated that GM-CSF Mϕs were significantly more responsive to IFN-λ3, as demonstrated by smear plot and Venn diagram of genes regulated above 2-fold (B) . GM-CSF Mϕ ISG induction was significantly stronger, as demonstrated by heat map of gene log 2-fold change (logFC) (C) . IFN-λ3 increased transcript abundance of numerous ISGs and transcription factors (TFs) in both Mϕ sets, as well as numerous chemokines, cytokines, and genes responsible for antigen (Ag) presentation and co-stimulation, particularly in GM-CSF Mϕs (D) . IFN-λ3 stimulated genes were confirmed by qPCR, using IFN-α differentiated Mϕs as a comparison (E) ( n = 8). Quantitative PCR data are representative of three independent experiments. Paired t -test * /# p
    Figure Legend Snippet: Monocyte differentiation with IFN-λ3 drives a pro-inflammatory macrophage phenotype. Monocytes were cultured with M- or GM-CSF ± IFN-λ3 for 7 days to examine the effect of IFN-λ3 on Mϕ differentiation, followed by phenotypic and functional characterization (A) . RNA sequencing ( n = 3/treatment) demonstrated that GM-CSF Mϕs were significantly more responsive to IFN-λ3, as demonstrated by smear plot and Venn diagram of genes regulated above 2-fold (B) . GM-CSF Mϕ ISG induction was significantly stronger, as demonstrated by heat map of gene log 2-fold change (logFC) (C) . IFN-λ3 increased transcript abundance of numerous ISGs and transcription factors (TFs) in both Mϕ sets, as well as numerous chemokines, cytokines, and genes responsible for antigen (Ag) presentation and co-stimulation, particularly in GM-CSF Mϕs (D) . IFN-λ3 stimulated genes were confirmed by qPCR, using IFN-α differentiated Mϕs as a comparison (E) ( n = 8). Quantitative PCR data are representative of three independent experiments. Paired t -test * /# p

    Techniques Used: Cell Culture, Functional Assay, RNA Sequencing Assay, Real-time Polymerase Chain Reaction

    4) Product Images from "Hypoxic resistance of KRAS mutant tumor cells to 3-Bromopyruvate is counteracted by Prima-1 and reversed by N-acetylcysteine"

    Article Title: Hypoxic resistance of KRAS mutant tumor cells to 3-Bromopyruvate is counteracted by Prima-1 and reversed by N-acetylcysteine

    Journal: BMC Cancer

    doi: 10.1186/s12885-016-2930-9

    a  3-BrPA potentiates Prima-1 toxicity against A549 cells in 5 mM glucose. A549 cells (4X10 3 ) were seeded in tissue culture 96 well plates in complete medium containing 20 mM glucose and 10% fetal bovine serum, then washed 3X with PBS and treated as indicated in each case, in medium supplemented with physiological 5 mM glucose, 2 mM glutamine and 5% dialyzed serum for 48 h. Relative proliferation /toxicity was assayed fluorometrically in octuplicate by the Alamar Blue method by quantitating the conversion of resazurin to fluorescent resorufin [  8 ]. This revealed that 50 μM Prima-1 cooperated with 3-BrPA rather than with CHC to suppress A549 cell growth.  b  Prima-1 decreases SLC2A1-GLUT1 in A549 cells. Sparse cells were seeded in 5 cm tissue culture plates (5  ×  10 5 cells per plate) in complete Dulbecco’s medium containing 20 mM glucose supplemented with 10% serum for 18 h, then washed 3X with PBS and treated as indicated in each case, in medium supplemented with physiological 5 mM glucose, 2 mM glutamine and 5% dialyzed serum whenever indicated (+) for 24 h. After RNA extraction with TRIZOL and quantification in a Qubit® 2.0 Fluorometer, cDNAs were prepared for end-point PCR analysis as indicated under Methods.essentially similar results were obtained in cells treated with Prima-1 in 5 mM glucose (not shown). Cells treated in parallel with those used for RNA analysis were used for GLUT1 protein immune blot [  40 ].  c  Prima-1 activates p21CDKN1A gene expression in A549 cells in 5 mM glucose. qPCR was used to determine relative expression of the p21CDK1N1 gene in control and treated cells, after RNA extraction, cDNA preparation and qPCR, as indicated under Methods. *denotes significance between treated cells relative to controls
    Figure Legend Snippet: a 3-BrPA potentiates Prima-1 toxicity against A549 cells in 5 mM glucose. A549 cells (4X10 3 ) were seeded in tissue culture 96 well plates in complete medium containing 20 mM glucose and 10% fetal bovine serum, then washed 3X with PBS and treated as indicated in each case, in medium supplemented with physiological 5 mM glucose, 2 mM glutamine and 5% dialyzed serum for 48 h. Relative proliferation /toxicity was assayed fluorometrically in octuplicate by the Alamar Blue method by quantitating the conversion of resazurin to fluorescent resorufin [ 8 ]. This revealed that 50 μM Prima-1 cooperated with 3-BrPA rather than with CHC to suppress A549 cell growth. b Prima-1 decreases SLC2A1-GLUT1 in A549 cells. Sparse cells were seeded in 5 cm tissue culture plates (5  ×  10 5 cells per plate) in complete Dulbecco’s medium containing 20 mM glucose supplemented with 10% serum for 18 h, then washed 3X with PBS and treated as indicated in each case, in medium supplemented with physiological 5 mM glucose, 2 mM glutamine and 5% dialyzed serum whenever indicated (+) for 24 h. After RNA extraction with TRIZOL and quantification in a Qubit® 2.0 Fluorometer, cDNAs were prepared for end-point PCR analysis as indicated under Methods.essentially similar results were obtained in cells treated with Prima-1 in 5 mM glucose (not shown). Cells treated in parallel with those used for RNA analysis were used for GLUT1 protein immune blot [ 40 ]. c Prima-1 activates p21CDKN1A gene expression in A549 cells in 5 mM glucose. qPCR was used to determine relative expression of the p21CDK1N1 gene in control and treated cells, after RNA extraction, cDNA preparation and qPCR, as indicated under Methods. *denotes significance between treated cells relative to controls

    Techniques Used: RNA Extraction, Polymerase Chain Reaction, Expressing, Real-time Polymerase Chain Reaction

    5) Product Images from "Identification of Balanced Chromosomal Rearrangements Previously Unknown Among Participants in the 1000 Genomes Project: Implications for Interpretation of Structural Variation in Genomes and the Future of Clinical Cytogenetics"

    Article Title: Identification of Balanced Chromosomal Rearrangements Previously Unknown Among Participants in the 1000 Genomes Project: Implications for Interpretation of Structural Variation in Genomes and the Future of Clinical Cytogenetics

    Journal: Genetics in medicine : official journal of the American College of Medical Genetics

    doi: 10.1038/gim.2017.170

    Gene disruption, cryptic deletions and potential disruption of interaction between promoter and enhancer by the breakpoints of balanced translocations Figures (A) , (B) and (C) NRXN3 disruption in 46,XX,t(9;14)(q34.2;q31.1) (HG02260). (A) and (B) Genomic locations of NRXN3 and RXRA are shown with breakpoints indicated by red dotted lines. (C) NRXN3 and RXRA expression for the four cases from the 1000 Genomes Project and for 13 reported EBV-B normal control cell lines (the GTEx project). Gene expression for NRXN3 and RXRA in HG02260 are indicated with red arrows. Figures (D) and (E) cryptic deletions involved at the breakpoints in translocation 46,XY,t(3;17)(q24;p13.3) (HG03729). Two cryptic deletions of seq[GRCh37/hg19] 3q24(143,817,430_143,822,651)×1 and seq[GRCh37/hg19] 17p13.3(2,910,366_2,914,751)×1 were detected by read-depth difference algorithm and were further confirmed by quantitative PCR. The deleted regions are shown in a yellow background with a red arrow while the normal copy-ratio (diploid) is shown in a blue background with a blue arrow. Two independent pairs of primers ( Supplementary Table 2 ) were used to perform qPCR in quintuplicate for validation of each deletion. The bars in cyan show the relative quantification of HG03729, while the bars in blue indicate the negative control. Figures (F) and (G) potential disruption of interaction between promoter and enhancer from rearrangement in 46,XY,t(16;17)(q23.1;q24.2) (NA18612) in H1-hESC. (F) Genes and the ChIP-seq data from the ENCODE Project are shown in terms of the genomic location. Each cell line with the ChIP-seq data ( i.e ., H3K4Me3 and H3K27Ac) 33 is labeled with a red arrow. Breakpoint in seq[GRCh37/hg19] 17q24.2(64,953,078_64,953,079) is shown by a green vertical line, while the candidate promoters and enhancers are indicated with orange and blue arrows, respectively. The region of potential enhancer in H1-hESC is highlighted in DNase I Hypersensitivity Clusters 34 in a blue rectangle (DHS region). The figure below is zoomed in on the potential enhancer region in H1-hESC. Enrichment of H3K4Me1 and absence of H3K4Me3 support a potential active enhancer in this region 33 , 39 , while enrichment of DNA-binding sequence motifs also indicates the candidate region of the interaction for regulatory elements 33 . (G) Gene expression level (Read Per Kilobase Million) of the four cases and 13 EBV-B normal control samples (GTEx project) 24 .
    Figure Legend Snippet: Gene disruption, cryptic deletions and potential disruption of interaction between promoter and enhancer by the breakpoints of balanced translocations Figures (A) , (B) and (C) NRXN3 disruption in 46,XX,t(9;14)(q34.2;q31.1) (HG02260). (A) and (B) Genomic locations of NRXN3 and RXRA are shown with breakpoints indicated by red dotted lines. (C) NRXN3 and RXRA expression for the four cases from the 1000 Genomes Project and for 13 reported EBV-B normal control cell lines (the GTEx project). Gene expression for NRXN3 and RXRA in HG02260 are indicated with red arrows. Figures (D) and (E) cryptic deletions involved at the breakpoints in translocation 46,XY,t(3;17)(q24;p13.3) (HG03729). Two cryptic deletions of seq[GRCh37/hg19] 3q24(143,817,430_143,822,651)×1 and seq[GRCh37/hg19] 17p13.3(2,910,366_2,914,751)×1 were detected by read-depth difference algorithm and were further confirmed by quantitative PCR. The deleted regions are shown in a yellow background with a red arrow while the normal copy-ratio (diploid) is shown in a blue background with a blue arrow. Two independent pairs of primers ( Supplementary Table 2 ) were used to perform qPCR in quintuplicate for validation of each deletion. The bars in cyan show the relative quantification of HG03729, while the bars in blue indicate the negative control. Figures (F) and (G) potential disruption of interaction between promoter and enhancer from rearrangement in 46,XY,t(16;17)(q23.1;q24.2) (NA18612) in H1-hESC. (F) Genes and the ChIP-seq data from the ENCODE Project are shown in terms of the genomic location. Each cell line with the ChIP-seq data ( i.e ., H3K4Me3 and H3K27Ac) 33 is labeled with a red arrow. Breakpoint in seq[GRCh37/hg19] 17q24.2(64,953,078_64,953,079) is shown by a green vertical line, while the candidate promoters and enhancers are indicated with orange and blue arrows, respectively. The region of potential enhancer in H1-hESC is highlighted in DNase I Hypersensitivity Clusters 34 in a blue rectangle (DHS region). The figure below is zoomed in on the potential enhancer region in H1-hESC. Enrichment of H3K4Me1 and absence of H3K4Me3 support a potential active enhancer in this region 33 , 39 , while enrichment of DNA-binding sequence motifs also indicates the candidate region of the interaction for regulatory elements 33 . (G) Gene expression level (Read Per Kilobase Million) of the four cases and 13 EBV-B normal control samples (GTEx project) 24 .

    Techniques Used: Expressing, Translocation Assay, Real-time Polymerase Chain Reaction, Negative Control, Chromatin Immunoprecipitation, Labeling, Binding Assay, Sequencing

    6) Product Images from "Comparative evaluation of rRNA depletion procedures for the improved analysis of bacterial biofilm and mixed pathogen culture transcriptomes"

    Article Title: Comparative evaluation of rRNA depletion procedures for the improved analysis of bacterial biofilm and mixed pathogen culture transcriptomes

    Journal: Scientific Reports

    doi: 10.1038/srep41114

    Quantitative and qualitative comparison of RNA recovered following rRNA depletion. A total of 4 μg of DNAse-treated RNA, isolated form 3-day-old P. aeruginosa PAO1 biofilms, was subjected to treatment with the Illumina Ribo-Zero rRNA Removal Kit (Bacteria), Ambion MICROBExpress™ Bacterial mRNA Enrichment Kit and the Life Technologies RiboMinus Transcriptome Isolation Kit, Bacteria. Following rRNA depletion and ethanol/acetate precipitation and resuspension in equal volumes of water, the RNA was assessed using Qubit fluorimetric quantitation with the Qubit RNA HS Assay Kit. Yields are reported as total RNA recovered ( A ) and as percentage of the input RNA ( B ). The RNA samples were also assessed using the Bioanalyzer RNA 6000 Pico kit. Representative electropherograms of ( C ) starting total RNA material and aliquots of the RNA samples that have been processed using the ( D ) MICROBExpress, ( E ) RiboMinus, or ( F ) Ribo-Zero kits are shown. Dashed lines indicate peaks corresponding to 16S and 23S RNA traces, which were detected in total RNA, MICROBExpress, and RiboMins samples. ( G ) The area of 16S and 23S rRNA peaks as percent of the total detected RNA was estimated, as determined using the 2100 Expert Software. RFU, relative fluorescence units. Experiments were repeated using three biological replicates.
    Figure Legend Snippet: Quantitative and qualitative comparison of RNA recovered following rRNA depletion. A total of 4 μg of DNAse-treated RNA, isolated form 3-day-old P. aeruginosa PAO1 biofilms, was subjected to treatment with the Illumina Ribo-Zero rRNA Removal Kit (Bacteria), Ambion MICROBExpress™ Bacterial mRNA Enrichment Kit and the Life Technologies RiboMinus Transcriptome Isolation Kit, Bacteria. Following rRNA depletion and ethanol/acetate precipitation and resuspension in equal volumes of water, the RNA was assessed using Qubit fluorimetric quantitation with the Qubit RNA HS Assay Kit. Yields are reported as total RNA recovered ( A ) and as percentage of the input RNA ( B ). The RNA samples were also assessed using the Bioanalyzer RNA 6000 Pico kit. Representative electropherograms of ( C ) starting total RNA material and aliquots of the RNA samples that have been processed using the ( D ) MICROBExpress, ( E ) RiboMinus, or ( F ) Ribo-Zero kits are shown. Dashed lines indicate peaks corresponding to 16S and 23S RNA traces, which were detected in total RNA, MICROBExpress, and RiboMins samples. ( G ) The area of 16S and 23S rRNA peaks as percent of the total detected RNA was estimated, as determined using the 2100 Expert Software. RFU, relative fluorescence units. Experiments were repeated using three biological replicates.

    Techniques Used: Isolation, Quantitation Assay, RNA HS Assay, Software, Fluorescence

    7) Product Images from "Hypoxic resistance of KRAS mutant tumor cells to 3-Bromopyruvate is counteracted by Prima-1 and reversed by N-acetylcysteine"

    Article Title: Hypoxic resistance of KRAS mutant tumor cells to 3-Bromopyruvate is counteracted by Prima-1 and reversed by N-acetylcysteine

    Journal: BMC Cancer

    doi: 10.1186/s12885-016-2930-9

    a 3-BrPA potentiates Prima-1 toxicity against A549 cells in 5 mM glucose. A549 cells (4X10 3 ) were seeded in tissue culture 96 well plates in complete medium containing 20 mM glucose and 10% fetal bovine serum, then washed 3X with PBS and treated as indicated in each case, in medium supplemented with physiological 5 mM glucose, 2 mM glutamine and 5% dialyzed serum for 48 h. Relative proliferation /toxicity was assayed fluorometrically in octuplicate by the Alamar Blue method by quantitating the conversion of resazurin to fluorescent resorufin [ 8 ]. This revealed that 50 μM Prima-1 cooperated with 3-BrPA rather than with CHC to suppress A549 cell growth. b Prima-1 decreases SLC2A1-GLUT1 in A549 cells. Sparse cells were seeded in 5 cm tissue culture plates (5 × 10 5 cells per plate) in complete Dulbecco’s medium containing 20 mM glucose supplemented with 10% serum for 18 h, then washed 3X with PBS and treated as indicated in each case, in medium supplemented with physiological 5 mM glucose, 2 mM glutamine and 5% dialyzed serum whenever indicated (+) for 24 h. After RNA extraction with TRIZOL and quantification in a Qubit® 2.0 Fluorometer, cDNAs were prepared for end-point PCR analysis as indicated under Methods.essentially similar results were obtained in cells treated with Prima-1 in 5 mM glucose (not shown). Cells treated in parallel with those used for RNA analysis were used for GLUT1 protein immune blot [ 40 ]. c Prima-1 activates p21CDKN1A gene expression in A549 cells in 5 mM glucose. qPCR was used to determine relative expression of the p21CDK1N1 gene in control and treated cells, after RNA extraction, cDNA preparation and qPCR, as indicated under Methods. *denotes significance between treated cells relative to controls
    Figure Legend Snippet: a 3-BrPA potentiates Prima-1 toxicity against A549 cells in 5 mM glucose. A549 cells (4X10 3 ) were seeded in tissue culture 96 well plates in complete medium containing 20 mM glucose and 10% fetal bovine serum, then washed 3X with PBS and treated as indicated in each case, in medium supplemented with physiological 5 mM glucose, 2 mM glutamine and 5% dialyzed serum for 48 h. Relative proliferation /toxicity was assayed fluorometrically in octuplicate by the Alamar Blue method by quantitating the conversion of resazurin to fluorescent resorufin [ 8 ]. This revealed that 50 μM Prima-1 cooperated with 3-BrPA rather than with CHC to suppress A549 cell growth. b Prima-1 decreases SLC2A1-GLUT1 in A549 cells. Sparse cells were seeded in 5 cm tissue culture plates (5 × 10 5 cells per plate) in complete Dulbecco’s medium containing 20 mM glucose supplemented with 10% serum for 18 h, then washed 3X with PBS and treated as indicated in each case, in medium supplemented with physiological 5 mM glucose, 2 mM glutamine and 5% dialyzed serum whenever indicated (+) for 24 h. After RNA extraction with TRIZOL and quantification in a Qubit® 2.0 Fluorometer, cDNAs were prepared for end-point PCR analysis as indicated under Methods.essentially similar results were obtained in cells treated with Prima-1 in 5 mM glucose (not shown). Cells treated in parallel with those used for RNA analysis were used for GLUT1 protein immune blot [ 40 ]. c Prima-1 activates p21CDKN1A gene expression in A549 cells in 5 mM glucose. qPCR was used to determine relative expression of the p21CDK1N1 gene in control and treated cells, after RNA extraction, cDNA preparation and qPCR, as indicated under Methods. *denotes significance between treated cells relative to controls

    Techniques Used: RNA Extraction, Polymerase Chain Reaction, Expressing, Real-time Polymerase Chain Reaction

    8) Product Images from "RUNX1 mitotically bookmarks target genes that are important for the mammary epithelial-to-mesenchymal transition"

    Article Title: RUNX1 mitotically bookmarks target genes that are important for the mammary epithelial-to-mesenchymal transition

    Journal: bioRxiv

    doi: 10.1101/511410

    RUNX1 bookmarks RNA Pol II-transcribed genes involved in maintenance of breast epithelial phenotype. A) Gene Set Enrichment (GSE) analysis from interrogating mitotically bookmarked genes (i.e. RUNX1 mitotically occupied) against Hallmark Gene sets from Molecular Signatures Database (MSigDB). The top 10 most significantly overlapping gene sets are shown from top to bottom. B) Scatter plot of genes identified to be up or down regulated in response to estradiol treatment, that are also bound by estrogen receptor α (ERα) and RUNX1 (empty circles, blue for downregulated and red for upregulated). Scatter plot also illustrates up or down regulated genes in response to estradiol treatment that are bound by ERα and mitotically bookmarked by RUNX1 (filled in circles, blue for downregulated and red for upregulated). C) Top panel: ChIP-Seq tracks of HES1 (left) and H2AFX (right) from asynchronous (top-red), mitotic (middle-green), and G1 (bottom-blue). Bottom panel: qRT-PCR data of HES1 (left) and H2AFX (right) in asynchronous MCF10A cells treated with either active (AI-14-91) or inactive (AI-4-88) inhibitors for 6hr, 12hr, 24hr and 48hr at 20μM. Expression of target genes were normalized relative to beta actin.
    Figure Legend Snippet: RUNX1 bookmarks RNA Pol II-transcribed genes involved in maintenance of breast epithelial phenotype. A) Gene Set Enrichment (GSE) analysis from interrogating mitotically bookmarked genes (i.e. RUNX1 mitotically occupied) against Hallmark Gene sets from Molecular Signatures Database (MSigDB). The top 10 most significantly overlapping gene sets are shown from top to bottom. B) Scatter plot of genes identified to be up or down regulated in response to estradiol treatment, that are also bound by estrogen receptor α (ERα) and RUNX1 (empty circles, blue for downregulated and red for upregulated). Scatter plot also illustrates up or down regulated genes in response to estradiol treatment that are bound by ERα and mitotically bookmarked by RUNX1 (filled in circles, blue for downregulated and red for upregulated). C) Top panel: ChIP-Seq tracks of HES1 (left) and H2AFX (right) from asynchronous (top-red), mitotic (middle-green), and G1 (bottom-blue). Bottom panel: qRT-PCR data of HES1 (left) and H2AFX (right) in asynchronous MCF10A cells treated with either active (AI-14-91) or inactive (AI-4-88) inhibitors for 6hr, 12hr, 24hr and 48hr at 20μM. Expression of target genes were normalized relative to beta actin.

    Techniques Used: Chromatin Immunoprecipitation, Quantitative RT-PCR, Expressing

    9) Product Images from "MP-12 virus containing the clone 13 deletion in the NSs gene prevents lethal disease when administered after Rift Valley fever virus infection in hamsters"

    Article Title: MP-12 virus containing the clone 13 deletion in the NSs gene prevents lethal disease when administered after Rift Valley fever virus infection in hamsters

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2015.00651

    Accumulation of RVFV S RNA and IFN-β mRNA in popliteal lymph nodes (LNs) of vaccinated hamsters. Hamsters were vaccinated s.c. with 1 × 10 5 PFU of rMP-12, rMP12-C13type or PBS (mock) in the left hind leg footpad. At 24 h post-vaccination, popliteal LNs were collected from the left and right hind legs. The RNA copies for (A) RVFV S RNA in the left and right popliteal LNs and for (B) IFN-β mRNA in the left popliteal LNs were measured by droplet digital PCR. Mean values are shown. Unique symbols represent the same animal across all parameters. The dashed line represents the limit of detection based on a mock-vaccinated control. * P
    Figure Legend Snippet: Accumulation of RVFV S RNA and IFN-β mRNA in popliteal lymph nodes (LNs) of vaccinated hamsters. Hamsters were vaccinated s.c. with 1 × 10 5 PFU of rMP-12, rMP12-C13type or PBS (mock) in the left hind leg footpad. At 24 h post-vaccination, popliteal LNs were collected from the left and right hind legs. The RNA copies for (A) RVFV S RNA in the left and right popliteal LNs and for (B) IFN-β mRNA in the left popliteal LNs were measured by droplet digital PCR. Mean values are shown. Unique symbols represent the same animal across all parameters. The dashed line represents the limit of detection based on a mock-vaccinated control. * P

    Techniques Used: Digital PCR

    10) Product Images from "The Genetic Basis of Host Preference and Resting Behavior in the Major African Malaria Vector, Anopheles arabiensis"

    Article Title: The Genetic Basis of Host Preference and Resting Behavior in the Major African Malaria Vector, Anopheles arabiensis

    Journal: PLoS Genetics

    doi: 10.1371/journal.pgen.1006303

    Genetic variation explained by the 2Rb and 3Ra inversions. a) Genetic structure was assessed using genome-wide SNP data for individual An . arabiensis females using a PCA analysis. Three discrete PCA clusters were observed. Red = human-fed and blue = cattle-fed. There is an enrichment of cattle-fed individuals in the middle PCA cluster (P
    Figure Legend Snippet: Genetic variation explained by the 2Rb and 3Ra inversions. a) Genetic structure was assessed using genome-wide SNP data for individual An . arabiensis females using a PCA analysis. Three discrete PCA clusters were observed. Red = human-fed and blue = cattle-fed. There is an enrichment of cattle-fed individuals in the middle PCA cluster (P

    Techniques Used: Genome Wide

    Relative host choice between villages. This figure describes the results of bloodmeal analysis of An . arabiensis collected from: Lupiro, Minepa, and Sagamaganga (SAGA). We detected multiple hosts in
    Figure Legend Snippet: Relative host choice between villages. This figure describes the results of bloodmeal analysis of An . arabiensis collected from: Lupiro, Minepa, and Sagamaganga (SAGA). We detected multiple hosts in

    Techniques Used:

    11) Product Images from "Direct CRISPR spacer acquisition from RNA by a natural reverse-transcriptase-Cas1 fusion protein"

    Article Title: Direct CRISPR spacer acquisition from RNA by a natural reverse-transcriptase-Cas1 fusion protein

    Journal: Science (New York, N.Y.)

    doi: 10.1126/science.aad4234

    cDNA synthesis using RNA ligated to CRISPR DNA
    Figure Legend Snippet: cDNA synthesis using RNA ligated to CRISPR DNA

    Techniques Used: CRISPR

    12) Product Images from "Functional DNA quantification guides accurate next-generation sequencing mutation detection in formalin-fixed, paraffin-embedded tumor biopsies"

    Article Title: Functional DNA quantification guides accurate next-generation sequencing mutation detection in formalin-fixed, paraffin-embedded tumor biopsies

    Journal: Genome Medicine

    doi: 10.1186/gm481

    FFPE DNA characterization by QFI-PCR and fluorescence-based assays from 165 tumor DNA samples. (A) Distribution of FFPE DNA quantification using QFI-PCR and the fluorescence-based Qubit dsDNA HS assay from 5 ng bulk DNA input as determined by NanoDrop spectrophotometry. A total of 27 samples were undetected by fluorescence assay (
    Figure Legend Snippet: FFPE DNA characterization by QFI-PCR and fluorescence-based assays from 165 tumor DNA samples. (A) Distribution of FFPE DNA quantification using QFI-PCR and the fluorescence-based Qubit dsDNA HS assay from 5 ng bulk DNA input as determined by NanoDrop spectrophotometry. A total of 27 samples were undetected by fluorescence assay (

    Techniques Used: Formalin-fixed Paraffin-Embedded, Polymerase Chain Reaction, Fluorescence, Spectrophotometry

    13) Product Images from "Direct oligonucleotide synthesis onto super-paramagnetic beads"

    Article Title: Direct oligonucleotide synthesis onto super-paramagnetic beads

    Journal: Journal of biotechnology

    doi: 10.1016/j.jbiotec.2013.08.006

    shows HPLC chromatograms of ( a ) Primer A′ (40 bp) synthesized on super-paramagnetic beads (SPMB)s, and ( b ) Primer A′ synthesized on CPG support as a control. Peak retention times are 4.817 and 4.819 minutes, respectively.
    Figure Legend Snippet: shows HPLC chromatograms of ( a ) Primer A′ (40 bp) synthesized on super-paramagnetic beads (SPMB)s, and ( b ) Primer A′ synthesized on CPG support as a control. Peak retention times are 4.817 and 4.819 minutes, respectively.

    Techniques Used: High Performance Liquid Chromatography, Synthesized

    14) Product Images from "Quantitative Viral Community DNA Analysis Reveals the Dominance of Single-Stranded DNA Viruses in Offshore Upper Bathyal Sediment from Tohoku, Japan"

    Article Title: Quantitative Viral Community DNA Analysis Reveals the Dominance of Single-Stranded DNA Viruses in Offshore Upper Bathyal Sediment from Tohoku, Japan

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2018.00075

    Principal component analysis (PCA) analysis of (A) ssDNA and (B) dsDNA viral sequence components in the relevant deep-sea sedimentary virome libraries from the off-Tohoku and other reference sites (Ogasawara Trench, Mariana Trench, and off Shimokita Peninsula) based on the taxonomic composition of viral communities. The respective off-Tohoku viral communities with the three different depths are shown in blue and the references are shown in red.
    Figure Legend Snippet: Principal component analysis (PCA) analysis of (A) ssDNA and (B) dsDNA viral sequence components in the relevant deep-sea sedimentary virome libraries from the off-Tohoku and other reference sites (Ogasawara Trench, Mariana Trench, and off Shimokita Peninsula) based on the taxonomic composition of viral communities. The respective off-Tohoku viral communities with the three different depths are shown in blue and the references are shown in red.

    Techniques Used: Sequencing

    15) Product Images from "RNA-sequencing reveals genome-wide long non-coding RNAs profiling associated with early development of diabetic nephropathy"

    Article Title: RNA-sequencing reveals genome-wide long non-coding RNAs profiling associated with early development of diabetic nephropathy

    Journal: Oncotarget

    doi: 10.18632/oncotarget.22405

    Transcriptome profiles of lncRNA genes associated with disease progression (A) Hierarchical clustering of lncRNA genes that are differentially expressed in at least one time point by comparison with that at W0; rows and columns correspond to samples and lncRNA genes, respectively. (B) Number of differentially expressed lncRNA genes at various time points compared with control (W0) with corrected P -value cutoff of 0.05. (C) Venn diagram of differentially expressed lncRNA genes at different times. (D) Gene ontology enrichment for week-specific and common differentially expressed lncRNA genes, presented as −log10 hypergeometric P -value for enrichment. (E) Validation for differentially expressed lncRNAs in PTCs (HK2)Left portion: Results from RNA-seq in mouse DN models ( * p
    Figure Legend Snippet: Transcriptome profiles of lncRNA genes associated with disease progression (A) Hierarchical clustering of lncRNA genes that are differentially expressed in at least one time point by comparison with that at W0; rows and columns correspond to samples and lncRNA genes, respectively. (B) Number of differentially expressed lncRNA genes at various time points compared with control (W0) with corrected P -value cutoff of 0.05. (C) Venn diagram of differentially expressed lncRNA genes at different times. (D) Gene ontology enrichment for week-specific and common differentially expressed lncRNA genes, presented as −log10 hypergeometric P -value for enrichment. (E) Validation for differentially expressed lncRNAs in PTCs (HK2)Left portion: Results from RNA-seq in mouse DN models ( * p

    Techniques Used: RNA Sequencing Assay

    16) Product Images from "Comparative evaluation of rRNA depletion procedures for the improved analysis of bacterial biofilm and mixed pathogen culture transcriptomes"

    Article Title: Comparative evaluation of rRNA depletion procedures for the improved analysis of bacterial biofilm and mixed pathogen culture transcriptomes

    Journal: Scientific Reports

    doi: 10.1038/srep41114

    Quantitative and qualitative comparison of RNA recovered following rRNA depletion. A total of 4 μg of DNAse-treated RNA, isolated form 3-day-old P. aeruginosa PAO1 biofilms, was subjected to treatment with the Illumina Ribo-Zero rRNA Removal Kit (Bacteria), Ambion MICROBExpress™ Bacterial mRNA Enrichment Kit and the Life Technologies RiboMinus Transcriptome Isolation Kit, Bacteria. Following rRNA depletion and ethanol/acetate precipitation and resuspension in equal volumes of water, the RNA was assessed using Qubit fluorimetric quantitation with the Qubit RNA HS Assay Kit. Yields are reported as total RNA recovered ( A ) and as percentage of the input RNA ( B ). The RNA samples were also assessed using the Bioanalyzer RNA 6000 Pico kit. Representative electropherograms of ( C ) starting total RNA material and aliquots of the RNA samples that have been processed using the ( D ) MICROBExpress, ( E ) RiboMinus, or ( F ) Ribo-Zero kits are shown. Dashed lines indicate peaks corresponding to 16S and 23S RNA traces, which were detected in total RNA, MICROBExpress, and RiboMins samples. ( G ) The area of 16S and 23S rRNA peaks as percent of the total detected RNA was estimated, as determined using the 2100 Expert Software. RFU, relative fluorescence units. Experiments were repeated using three biological replicates.
    Figure Legend Snippet: Quantitative and qualitative comparison of RNA recovered following rRNA depletion. A total of 4 μg of DNAse-treated RNA, isolated form 3-day-old P. aeruginosa PAO1 biofilms, was subjected to treatment with the Illumina Ribo-Zero rRNA Removal Kit (Bacteria), Ambion MICROBExpress™ Bacterial mRNA Enrichment Kit and the Life Technologies RiboMinus Transcriptome Isolation Kit, Bacteria. Following rRNA depletion and ethanol/acetate precipitation and resuspension in equal volumes of water, the RNA was assessed using Qubit fluorimetric quantitation with the Qubit RNA HS Assay Kit. Yields are reported as total RNA recovered ( A ) and as percentage of the input RNA ( B ). The RNA samples were also assessed using the Bioanalyzer RNA 6000 Pico kit. Representative electropherograms of ( C ) starting total RNA material and aliquots of the RNA samples that have been processed using the ( D ) MICROBExpress, ( E ) RiboMinus, or ( F ) Ribo-Zero kits are shown. Dashed lines indicate peaks corresponding to 16S and 23S RNA traces, which were detected in total RNA, MICROBExpress, and RiboMins samples. ( G ) The area of 16S and 23S rRNA peaks as percent of the total detected RNA was estimated, as determined using the 2100 Expert Software. RFU, relative fluorescence units. Experiments were repeated using three biological replicates.

    Techniques Used: Isolation, Quantitation Assay, RNA HS Assay, Software, Fluorescence

    17) Product Images from "Comparative evaluation of rRNA depletion procedures for the improved analysis of bacterial biofilm and mixed pathogen culture transcriptomes"

    Article Title: Comparative evaluation of rRNA depletion procedures for the improved analysis of bacterial biofilm and mixed pathogen culture transcriptomes

    Journal: Scientific Reports

    doi: 10.1038/srep41114

    Quantitative and qualitative comparison of RNA recovered following rRNA depletion. A total of 4 μg of DNAse-treated RNA, isolated form 3-day-old P. aeruginosa PAO1 biofilms, was subjected to treatment with the Illumina Ribo-Zero rRNA Removal Kit (Bacteria), Ambion MICROBExpress™ Bacterial mRNA Enrichment Kit and the Life Technologies RiboMinus Transcriptome Isolation Kit, Bacteria. Following rRNA depletion and ethanol/acetate precipitation and resuspension in equal volumes of water, the RNA was assessed using Qubit fluorimetric quantitation with the Qubit RNA HS Assay Kit. Yields are reported as total RNA recovered ( A ) and as percentage of the input RNA ( B ). The RNA samples were also assessed using the Bioanalyzer RNA 6000 Pico kit. Representative electropherograms of ( C ) starting total RNA material and aliquots of the RNA samples that have been processed using the ( D ) MICROBExpress, ( E ) RiboMinus, or ( F ) Ribo-Zero kits are shown. Dashed lines indicate peaks corresponding to 16S and 23S RNA traces, which were detected in total RNA, MICROBExpress, and RiboMins samples. ( G ) The area of 16S and 23S rRNA peaks as percent of the total detected RNA was estimated, as determined using the 2100 Expert Software. RFU, relative fluorescence units. Experiments were repeated using three biological replicates.
    Figure Legend Snippet: Quantitative and qualitative comparison of RNA recovered following rRNA depletion. A total of 4 μg of DNAse-treated RNA, isolated form 3-day-old P. aeruginosa PAO1 biofilms, was subjected to treatment with the Illumina Ribo-Zero rRNA Removal Kit (Bacteria), Ambion MICROBExpress™ Bacterial mRNA Enrichment Kit and the Life Technologies RiboMinus Transcriptome Isolation Kit, Bacteria. Following rRNA depletion and ethanol/acetate precipitation and resuspension in equal volumes of water, the RNA was assessed using Qubit fluorimetric quantitation with the Qubit RNA HS Assay Kit. Yields are reported as total RNA recovered ( A ) and as percentage of the input RNA ( B ). The RNA samples were also assessed using the Bioanalyzer RNA 6000 Pico kit. Representative electropherograms of ( C ) starting total RNA material and aliquots of the RNA samples that have been processed using the ( D ) MICROBExpress, ( E ) RiboMinus, or ( F ) Ribo-Zero kits are shown. Dashed lines indicate peaks corresponding to 16S and 23S RNA traces, which were detected in total RNA, MICROBExpress, and RiboMins samples. ( G ) The area of 16S and 23S rRNA peaks as percent of the total detected RNA was estimated, as determined using the 2100 Expert Software. RFU, relative fluorescence units. Experiments were repeated using three biological replicates.

    Techniques Used: Isolation, Quantitation Assay, RNA HS Assay, Software, Fluorescence

    18) Product Images from "Early Recovery of Salmonella from Food Using a 6-Hour Non-selective Pre-enrichment and Reformulation of Tetrathionate Broth"

    Article Title: Early Recovery of Salmonella from Food Using a 6-Hour Non-selective Pre-enrichment and Reformulation of Tetrathionate Broth

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2016.02103

    Proportional abundances of bacteria in Tetrathionate (TT) and Rappaport–Vassiliadis (RV) selective enrichments . Proportional abundances of Salmonella and other family members from TT and RV selective enrichments were estimated using 16S rRNA gene sequencing. sRV and sTT (5 and 6-h RV and TT), 24 RV and 24TT (24-h RV and TT), ∗ significant increase in sRV and sTT B verses 24RV or 24TT B , ∗∗ significant decrease in sTT compared to 24 TT, and ∗∗∗ significant increase in 24RV versus 24TT B .
    Figure Legend Snippet: Proportional abundances of bacteria in Tetrathionate (TT) and Rappaport–Vassiliadis (RV) selective enrichments . Proportional abundances of Salmonella and other family members from TT and RV selective enrichments were estimated using 16S rRNA gene sequencing. sRV and sTT (5 and 6-h RV and TT), 24 RV and 24TT (24-h RV and TT), ∗ significant increase in sRV and sTT B verses 24RV or 24TT B , ∗∗ significant decrease in sTT compared to 24 TT, and ∗∗∗ significant increase in 24RV versus 24TT B .

    Techniques Used: Sequencing

    Workflow for food processing and sample collection. All experiments used Salmonella positive and negative food samples that were aged and processed for Salmonella detection using two workflows. Non-selective enrichment samples were collected at 5, and 6-h (workflow 1, green) and 24-h (workflow 2, yellow) for Salmonella enumeration, inoculation of selective TT and RV enrichments and 16S rRNA gene sequencing. Daily sample processing steps were performed as outline on Day 1 (Blue), Day 2 (Pink), Day 3 (Gray), and Day 4 (Orange).
    Figure Legend Snippet: Workflow for food processing and sample collection. All experiments used Salmonella positive and negative food samples that were aged and processed for Salmonella detection using two workflows. Non-selective enrichment samples were collected at 5, and 6-h (workflow 1, green) and 24-h (workflow 2, yellow) for Salmonella enumeration, inoculation of selective TT and RV enrichments and 16S rRNA gene sequencing. Daily sample processing steps were performed as outline on Day 1 (Blue), Day 2 (Pink), Day 3 (Gray), and Day 4 (Orange).

    Techniques Used: Sequencing

    19) Product Images from "MP-12 virus containing the clone 13 deletion in the NSs gene prevents lethal disease when administered after Rift Valley fever virus infection in hamsters"

    Article Title: MP-12 virus containing the clone 13 deletion in the NSs gene prevents lethal disease when administered after Rift Valley fever virus infection in hamsters

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2015.00651

    Accumulation of RVFV S RNA and IFN-β mRNA in popliteal lymph nodes (LNs) of vaccinated hamsters. Hamsters were vaccinated s.c. with 1 × 10 5 PFU of rMP-12, rMP12-C13type or PBS (mock) in the left hind leg footpad. At 24 h post-vaccination, popliteal LNs were collected from the left and right hind legs. The RNA copies for (A) RVFV S RNA in the left and right popliteal LNs and for (B) IFN-β mRNA in the left popliteal LNs were measured by droplet digital PCR. Mean values are shown. Unique symbols represent the same animal across all parameters. The dashed line represents the limit of detection based on a mock-vaccinated control. * P
    Figure Legend Snippet: Accumulation of RVFV S RNA and IFN-β mRNA in popliteal lymph nodes (LNs) of vaccinated hamsters. Hamsters were vaccinated s.c. with 1 × 10 5 PFU of rMP-12, rMP12-C13type or PBS (mock) in the left hind leg footpad. At 24 h post-vaccination, popliteal LNs were collected from the left and right hind legs. The RNA copies for (A) RVFV S RNA in the left and right popliteal LNs and for (B) IFN-β mRNA in the left popliteal LNs were measured by droplet digital PCR. Mean values are shown. Unique symbols represent the same animal across all parameters. The dashed line represents the limit of detection based on a mock-vaccinated control. * P

    Techniques Used: Digital PCR

    20) Product Images from "Genomics Reveals a Unique Clone of Burkholderia cenocepacia Harboring an Actively Excising Novel Genomic Island"

    Article Title: Genomics Reveals a Unique Clone of Burkholderia cenocepacia Harboring an Actively Excising Novel Genomic Island

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2017.00590

    Circular representation of Burkholderia cenocepacia genomes to mark the genomic location of BcenGI15. (A) Genomes of various B. cenocepacia (BC) strains are visualized in the circular form by mapping on strain IPCU-A as a reference. One ring represents each genome; yellow rings represent the nosocomial isolates sequenced in this study, and all other rings are of the genomes of other B. cenocepacia (BC) strains available at NCBI database. The GC content (%) of IPCU-A is shown by a ragged inner circle in black. The novel genomic island (BcenGI15) associated with nosocomial isolates is compared across all the genomes. (B) Artemis comparison of contig 81 of IPCU-A containing BcenGI15 with reference genome B. cenocepacia J2315. Red color shows the region of forward matches. The GC content (%) graph is drawn in black to show the variability.
    Figure Legend Snippet: Circular representation of Burkholderia cenocepacia genomes to mark the genomic location of BcenGI15. (A) Genomes of various B. cenocepacia (BC) strains are visualized in the circular form by mapping on strain IPCU-A as a reference. One ring represents each genome; yellow rings represent the nosocomial isolates sequenced in this study, and all other rings are of the genomes of other B. cenocepacia (BC) strains available at NCBI database. The GC content (%) of IPCU-A is shown by a ragged inner circle in black. The novel genomic island (BcenGI15) associated with nosocomial isolates is compared across all the genomes. (B) Artemis comparison of contig 81 of IPCU-A containing BcenGI15 with reference genome B. cenocepacia J2315. Red color shows the region of forward matches. The GC content (%) graph is drawn in black to show the variability.

    Techniques Used:

    21) Product Images from "Sybr Green- and TaqMan-Based Quantitative PCR Approaches Allow Assessment of the Abundance and Relative Distribution of Frankia Clusters in Soils"

    Article Title: Sybr Green- and TaqMan-Based Quantitative PCR Approaches Allow Assessment of the Abundance and Relative Distribution of Frankia Clusters in Soils

    Journal: Applied and Environmental Microbiology

    doi: 10.1128/AEM.02833-16

    Sybr Green- and TaqMan-based qPCR quantification of amplicons from representative pure cultures or endophytes of Frankia clusters 1, 2, 3, and 4 and of subgroups a, b, and c within cluster 1. Individual amplicons or mixtures of amplicons were quantified
    Figure Legend Snippet: Sybr Green- and TaqMan-based qPCR quantification of amplicons from representative pure cultures or endophytes of Frankia clusters 1, 2, 3, and 4 and of subgroups a, b, and c within cluster 1. Individual amplicons or mixtures of amplicons were quantified

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

    22) Product Images from "Manipulation of both virus- and cell-specific factors is required for robust transient replication of a hepatitis C virus genotype 3a sub-genomic replicon"

    Article Title: Manipulation of both virus- and cell-specific factors is required for robust transient replication of a hepatitis C virus genotype 3a sub-genomic replicon

    Journal: The Journal of General Virology

    doi: 10.1099/jgv.0.000932

    Comparison of transient and stable replication of S52 SGR. (a) Cells harbouring SGR-Feo-S52, or SGR-Feo-S52 selected with DCV, were seeded at a density of 8×10 3 per well of a 96-well plate and harvested for luciferase assay after 72 h. Luciferase activity is presented as absolute values. (b) Two micrograms of RNA transcribed from SGR-neo-S52 or SGR-neo-S52(Y93H) were electroporated into Huh7.5 cells and seeded at a density of 2×10 5 cells per well of a 6-well plate. Cells were selected with 0.5 mg ml −1 G418 from 48 hpe for three weeks before being fixed and stained with 0.5 % crystal violet in 10 % formalin and manually counted. (c) Two micrograms of RNA transcribed from S52(A1672V) or S52(A1672V) Y93H was electroporated into VSEC cells and 1×10 4 cells were harvested for luciferase assay at 96 hpe. *** P ≤0.001, **** P ≤0.0001, ns: not significant.
    Figure Legend Snippet: Comparison of transient and stable replication of S52 SGR. (a) Cells harbouring SGR-Feo-S52, or SGR-Feo-S52 selected with DCV, were seeded at a density of 8×10 3 per well of a 96-well plate and harvested for luciferase assay after 72 h. Luciferase activity is presented as absolute values. (b) Two micrograms of RNA transcribed from SGR-neo-S52 or SGR-neo-S52(Y93H) were electroporated into Huh7.5 cells and seeded at a density of 2×10 5 cells per well of a 6-well plate. Cells were selected with 0.5 mg ml −1 G418 from 48 hpe for three weeks before being fixed and stained with 0.5 % crystal violet in 10 % formalin and manually counted. (c) Two micrograms of RNA transcribed from S52(A1672V) or S52(A1672V) Y93H was electroporated into VSEC cells and 1×10 4 cells were harvested for luciferase assay at 96 hpe. *** P ≤0.001, **** P ≤0.0001, ns: not significant.

    Techniques Used: Luciferase, Activity Assay, Staining

    Culture-adaptive substitutions identified following selection of S52 SGR-harbouring cells. Location of putative culture-adaptive substitutions on three-dimensional structures of NS3 or NS4A. In (a) K1406 is shown in green within NS3 helicase. Proposed spring helix shown in yellow, nucleic acid binding cleft in blue and ATP-binding cleft shown in red. In (b) H1685 is shown in green within the NS3-binding region of the NS4A peptide (red), and the NS3 protease-active site shown in blue. A1672 is within the hydrophobic N terminus (not shown on crystal structure). (c) Transient replication of the S52 SGR culture-adaptive substitutions. The indicated substitutions were cloned back into S52 SGR, and 2 µg RNA transcripts were electroporated into Huh7.5 cells and harvested for luciferase assay. Luciferase activity is presented as absolute values. Error bars show the standard error of the mean of three experimental repeats.
    Figure Legend Snippet: Culture-adaptive substitutions identified following selection of S52 SGR-harbouring cells. Location of putative culture-adaptive substitutions on three-dimensional structures of NS3 or NS4A. In (a) K1406 is shown in green within NS3 helicase. Proposed spring helix shown in yellow, nucleic acid binding cleft in blue and ATP-binding cleft shown in red. In (b) H1685 is shown in green within the NS3-binding region of the NS4A peptide (red), and the NS3 protease-active site shown in blue. A1672 is within the hydrophobic N terminus (not shown on crystal structure). (c) Transient replication of the S52 SGR culture-adaptive substitutions. The indicated substitutions were cloned back into S52 SGR, and 2 µg RNA transcripts were electroporated into Huh7.5 cells and harvested for luciferase assay. Luciferase activity is presented as absolute values. Error bars show the standard error of the mean of three experimental repeats.

    Techniques Used: Selection, Binding Assay, Clone Assay, Luciferase, Activity Assay

    Assay for replication of S52 SGR. (a) Transient replication of the S52 SGR (AII culture adapted variant) [ 16 ] compared to either wild-type or GND mutant JFH-1 (GT2a). Two micrograms of the indicated RNA transcripts were electroporated into Huh7.5 cells and harvested for luciferase assay at the indicated time points. Relative luciferase units are expressed as the ratio to 4 hpe. Error bars show standard error of the mean of three experimental repeats. (b) S52 SGR RNA was electroporated into Huh7.5 cells and selected with 0.5 mg ml −1 G418 from 48 hpe. Surviving colonies were pooled into a polyclonal population of SGR-harbouring cells. Luciferase activity was measured in 8×10 3 cells and presented as absolute values compared to Con1- and JFH-1 SGR-harbouring cell lines. (c) Western blot analysis of NS5A expression in JFH-1- or S52 SGR-harbouring cells. (d) S52- and JFH-1 SGR-harbouring cells were immunostained for NS5A (green) using a sheep polyclonal anti-NS5A serum and nuclei using DAPI. ** P ≤0.01, **** P ≤0.0001.
    Figure Legend Snippet: Assay for replication of S52 SGR. (a) Transient replication of the S52 SGR (AII culture adapted variant) [ 16 ] compared to either wild-type or GND mutant JFH-1 (GT2a). Two micrograms of the indicated RNA transcripts were electroporated into Huh7.5 cells and harvested for luciferase assay at the indicated time points. Relative luciferase units are expressed as the ratio to 4 hpe. Error bars show standard error of the mean of three experimental repeats. (b) S52 SGR RNA was electroporated into Huh7.5 cells and selected with 0.5 mg ml −1 G418 from 48 hpe. Surviving colonies were pooled into a polyclonal population of SGR-harbouring cells. Luciferase activity was measured in 8×10 3 cells and presented as absolute values compared to Con1- and JFH-1 SGR-harbouring cell lines. (c) Western blot analysis of NS5A expression in JFH-1- or S52 SGR-harbouring cells. (d) S52- and JFH-1 SGR-harbouring cells were immunostained for NS5A (green) using a sheep polyclonal anti-NS5A serum and nuclei using DAPI. ** P ≤0.01, **** P ≤0.0001.

    Techniques Used: Variant Assay, Mutagenesis, Luciferase, Activity Assay, Western Blot, Expressing

    Enhancement of SGR replication using low-CpG luciferase, PIV-5 V protein and SEC14L2. (a) Confirmation of PIV-5 V or SEC14L2 expression by western blot or RT-PCR, respectively. (b) Huh7.5 or Huh7.5 V cells were transfected with an ISRE-luc plasmid and treated with IFN-α for 6 or 12 h prior to harvest for luciferase assay. Two micrograms of the indicated SGR RNA transcripts were electroporated into Huh7.5 (c), Huh7.5 V (d), Huh7.5 SEC14L2 (e) and Huh7.5 V+SEC14 L2 (termed VSEC cells) (f). (g) For comparative purposes all data were combined on to a single graph. Cells were harvested for luciferase assay at the indicated time points. Relative luciferase units are expressed as the ratio to 4 hpe. Error bars represent the standard error of the mean of four experimental repeats. ** P ≤0.01, *** P ≤0.001, **** P ≤0.0001.
    Figure Legend Snippet: Enhancement of SGR replication using low-CpG luciferase, PIV-5 V protein and SEC14L2. (a) Confirmation of PIV-5 V or SEC14L2 expression by western blot or RT-PCR, respectively. (b) Huh7.5 or Huh7.5 V cells were transfected with an ISRE-luc plasmid and treated with IFN-α for 6 or 12 h prior to harvest for luciferase assay. Two micrograms of the indicated SGR RNA transcripts were electroporated into Huh7.5 (c), Huh7.5 V (d), Huh7.5 SEC14L2 (e) and Huh7.5 V+SEC14 L2 (termed VSEC cells) (f). (g) For comparative purposes all data were combined on to a single graph. Cells were harvested for luciferase assay at the indicated time points. Relative luciferase units are expressed as the ratio to 4 hpe. Error bars represent the standard error of the mean of four experimental repeats. ** P ≤0.01, *** P ≤0.001, **** P ≤0.0001.

    Techniques Used: Luciferase, Expressing, Western Blot, Reverse Transcription Polymerase Chain Reaction, Transfection, Plasmid Preparation

    23) Product Images from "Rapid molecular genetic diagnosis of hypertrophic cardiomyopathy by semiconductor sequencing"

    Article Title: Rapid molecular genetic diagnosis of hypertrophic cardiomyopathy by semiconductor sequencing

    Journal: Journal of Translational Medicine

    doi: 10.1186/1479-5876-12-173

    Semiconductor sequencing flow diagram. Amplicons of 30 HCM genes are amplified by using 20 ng input genomic DNA and two primer pools. The about 200 bp amplicons are end-polished, barcodes and adapters ligated, purified to become prepared libraries. Emulsion PCR is conducted by Ion One Touch to generate ISPs for semiconductor sequencing. ISP indicates Ion Sphere Particles.
    Figure Legend Snippet: Semiconductor sequencing flow diagram. Amplicons of 30 HCM genes are amplified by using 20 ng input genomic DNA and two primer pools. The about 200 bp amplicons are end-polished, barcodes and adapters ligated, purified to become prepared libraries. Emulsion PCR is conducted by Ion One Touch to generate ISPs for semiconductor sequencing. ISP indicates Ion Sphere Particles.

    Techniques Used: Sequencing, Flow Cytometry, Amplification, Purification, Polymerase Chain Reaction

    24) Product Images from "Reduced PRC2 function alters male germline epigenetic programming and paternal inheritance"

    Article Title: Reduced PRC2 function alters male germline epigenetic programming and paternal inheritance

    Journal: BMC Biology

    doi: 10.1186/s12915-018-0569-5

    Breeding and experimental plan to assess epigenetic inheritance. Wild-type ( wt/wt ) females that had never been exposed to the Eed hypomorphic mutation were mated to either a homozygous ( Eed hypo/hypo ), b heterozygous ( Eed hypo/wt ) males ( n = 3 littermate pairs for each genotype) or c wild-type ( Eed wt/wt , n = 2) males. Sperm in heterozygous and wild-type males develop with at least one fully functional copy of Eed ( b and c ), while sperm in homozygous hypomorphic males ( a ) develop without normally functioning Eed and are expected to contain altered epigenetic patterning. All oocytes are wild-type. d–g Stage- and size-matched E8.5-day embryos were collected for transcriptional analysis. Comparisons between heterozygous embryos ( d and e ) and between wild-type embryos ( f and g ) identified genes that were misregulated due to non-genetic differences inherited from the sperm
    Figure Legend Snippet: Breeding and experimental plan to assess epigenetic inheritance. Wild-type ( wt/wt ) females that had never been exposed to the Eed hypomorphic mutation were mated to either a homozygous ( Eed hypo/hypo ), b heterozygous ( Eed hypo/wt ) males ( n = 3 littermate pairs for each genotype) or c wild-type ( Eed wt/wt , n = 2) males. Sperm in heterozygous and wild-type males develop with at least one fully functional copy of Eed ( b and c ), while sperm in homozygous hypomorphic males ( a ) develop without normally functioning Eed and are expected to contain altered epigenetic patterning. All oocytes are wild-type. d–g Stage- and size-matched E8.5-day embryos were collected for transcriptional analysis. Comparisons between heterozygous embryos ( d and e ) and between wild-type embryos ( f and g ) identified genes that were misregulated due to non-genetic differences inherited from the sperm

    Techniques Used: Mutagenesis, Functional Assay

    25) Product Images from "Novel insights into neuroinflammation: bacterial lipopolysaccharide, tumor necrosis factor α, and Ureaplasma species differentially modulate atypical chemokine receptor 3 responses in human brain microvascular endothelial cells"

    Article Title: Novel insights into neuroinflammation: bacterial lipopolysaccharide, tumor necrosis factor α, and Ureaplasma species differentially modulate atypical chemokine receptor 3 responses in human brain microvascular endothelial cells

    Journal: Journal of Neuroinflammation

    doi: 10.1186/s12974-018-1170-0

    ACKR3 mRNA and protein expression in HBMEC after co-stimulation with LPS and Ureaplasma spp. ACKR3 mRNA expression was assessed using qRT-PCR ( a ) and RNA sequencing ( b ). Protein expression was determined by flow cytometry, with ( c ) representing the percentage of ACKR3- and CD31-positive HBMEC, and ( d ) depicting representative dot plots, both after a stimulation period of 48 h. Dot plots were gated for vital cells. Data are shown as mean ± SD ( p values after logarithmic transformation, °° p
    Figure Legend Snippet: ACKR3 mRNA and protein expression in HBMEC after co-stimulation with LPS and Ureaplasma spp. ACKR3 mRNA expression was assessed using qRT-PCR ( a ) and RNA sequencing ( b ). Protein expression was determined by flow cytometry, with ( c ) representing the percentage of ACKR3- and CD31-positive HBMEC, and ( d ) depicting representative dot plots, both after a stimulation period of 48 h. Dot plots were gated for vital cells. Data are shown as mean ± SD ( p values after logarithmic transformation, °° p

    Techniques Used: Expressing, Quantitative RT-PCR, RNA Sequencing Assay, Flow Cytometry, Cytometry, Transformation Assay

    ACKR3 mRNA and protein expression in HBMEC after stimulation with LPS, TNF-α, or Ureaplasma spp. Levels of mRNA were assessed by qRT-PCR ( a ) and RNA sequencing ( b ). Flow cytometry was used to determine protein expression, events were gated for vital cells, with CD31 functioning as endothelial marker. For a stimulation period of 48 h, ( c ) shows the percentage of ACKR3- and CD31-positive cells, whereas dot plots of one representative experiment are given in ( d ). Data are presented as mean ± SD and significances are reported for comparisons vs. control and vs. broth ( p values after logarithmic transformation, * p
    Figure Legend Snippet: ACKR3 mRNA and protein expression in HBMEC after stimulation with LPS, TNF-α, or Ureaplasma spp. Levels of mRNA were assessed by qRT-PCR ( a ) and RNA sequencing ( b ). Flow cytometry was used to determine protein expression, events were gated for vital cells, with CD31 functioning as endothelial marker. For a stimulation period of 48 h, ( c ) shows the percentage of ACKR3- and CD31-positive cells, whereas dot plots of one representative experiment are given in ( d ). Data are presented as mean ± SD and significances are reported for comparisons vs. control and vs. broth ( p values after logarithmic transformation, * p

    Techniques Used: Expressing, Quantitative RT-PCR, RNA Sequencing Assay, Flow Cytometry, Cytometry, Marker, Transformation Assay

    26) Product Images from "A Portable, Shock-Proof, Surface-Heated Droplet PCR System for Escherichia coli Detection"

    Article Title: A Portable, Shock-Proof, Surface-Heated Droplet PCR System for Escherichia coli Detection

    Journal: Biosensors & bioelectronics

    doi: 10.1016/j.bios.2015.06.026

    (A) Gel electropherogram showing results from the positive control experiment using 2.6 ng of genomic DNA (equivalent to 5.2×10 5 genomic copies) extracted from E. coli K12 and thermocycled for 30 cycles, with thermal cycle times of 30 s, on the surface-heated droplet PCR device. The genomic DNA was quantified by a Qubit 2.0 fluorimeter. The 196 bp product band is at the expected location, and there is no band observed for the no template control (NTC) sample. (B) Gel electropherogram showing the result of the dilution test. Genomic DNA in the range of 2.6 ng to 5.2 pg (5.2×10 5 − 10 3 genomic copies) was thermocycled for 30 cycles, with thermocycle times of 30 s, on the surface-heated droplet PCR device. The PCR with the lowest DNA content (5.2 pg or approximately 10 3 genomic copies) produced a visible band.
    Figure Legend Snippet: (A) Gel electropherogram showing results from the positive control experiment using 2.6 ng of genomic DNA (equivalent to 5.2×10 5 genomic copies) extracted from E. coli K12 and thermocycled for 30 cycles, with thermal cycle times of 30 s, on the surface-heated droplet PCR device. The genomic DNA was quantified by a Qubit 2.0 fluorimeter. The 196 bp product band is at the expected location, and there is no band observed for the no template control (NTC) sample. (B) Gel electropherogram showing the result of the dilution test. Genomic DNA in the range of 2.6 ng to 5.2 pg (5.2×10 5 − 10 3 genomic copies) was thermocycled for 30 cycles, with thermocycle times of 30 s, on the surface-heated droplet PCR device. The PCR with the lowest DNA content (5.2 pg or approximately 10 3 genomic copies) produced a visible band.

    Techniques Used: Positive Control, Polymerase Chain Reaction, Produced

    27) Product Images from "A CD8α− Subset of CD4+SLAMF7+ Cytotoxic T Cells is Expanded in Patients with IgG4-Related Disease and Decreases following Glucocorticoid Treatment"

    Article Title: A CD8α− Subset of CD4+SLAMF7+ Cytotoxic T Cells is Expanded in Patients with IgG4-Related Disease and Decreases following Glucocorticoid Treatment

    Journal: Arthritis & rheumatology (Hoboken, N.J.)

    doi: 10.1002/art.40469

    Expression of cytolytic molecules on CD8α − and CD8α low CD4 + SLAMF7 + T EM cells (A) Contour plots showing different frequencies of granzyme A and perforin positive CD8α − and CD8α low CD4 + T EM SLAMF7 + CTLs in a patient with IgG4-RD representative of n=10 independent experiments. ( B ) Immunofluorescence on bulk sorted CD8α − CD4 + T EM SLAMF7 + CTLs showing granzyme A production. ( C ) Histograms showing the expression of granzyme A and perforin on CD8α − and CD8α low CD4 + T EM SLAMF7 + CTLs in in a patient with IgG4-RD representative of n=10 independent experiments. *Gated on CD8+ T lymphocytes. ( D ) Expression level of CD8α on CD8α low CD4 + SLAMF7 + CTLs in a representative patient with IgG4-RD and in a representative healthy control (histograms), and in the study cohort presented as Median Fluorescence Intensity (MFI) (* gated on CD8+ T lymphocytes; ** p
    Figure Legend Snippet: Expression of cytolytic molecules on CD8α − and CD8α low CD4 + SLAMF7 + T EM cells (A) Contour plots showing different frequencies of granzyme A and perforin positive CD8α − and CD8α low CD4 + T EM SLAMF7 + CTLs in a patient with IgG4-RD representative of n=10 independent experiments. ( B ) Immunofluorescence on bulk sorted CD8α − CD4 + T EM SLAMF7 + CTLs showing granzyme A production. ( C ) Histograms showing the expression of granzyme A and perforin on CD8α − and CD8α low CD4 + T EM SLAMF7 + CTLs in in a patient with IgG4-RD representative of n=10 independent experiments. *Gated on CD8+ T lymphocytes. ( D ) Expression level of CD8α on CD8α low CD4 + SLAMF7 + CTLs in a representative patient with IgG4-RD and in a representative healthy control (histograms), and in the study cohort presented as Median Fluorescence Intensity (MFI) (* gated on CD8+ T lymphocytes; ** p

    Techniques Used: Expressing, Immunofluorescence, Fluorescence

    Clinical improvement induced by glucocorticoid treatment correlates with contraction of clonally expanded circulating CD8α − CD4 + SLAMF7 + T EM cells ( A ) Retrieval frequency of CDR3 amino acid sequences of TCRα and β chain from circulating CD8α-CD4 + SLAMF7 + T EM cells before and after treatment and in the pancreas of Patient #1, and in the blood, lacrimal gland, and lung tissues of Patient #5 show an oligoclonal pattern on Next-Generation Sequencing analysis. Circulating CD4 + T EM cells from a healthy donor are polyclonal. Red bars indicate the most frequently encountered sequences; violet bars represent the 10th most frequently encountered sequences; grey bars represent overall remaining CDR3 sequences. The higher the grey bar, the more polyclonal is the sample. Patient #1 also shows an increased polyclonality of the TCRα and β chain repertoire after treatment. ( B ) Ten most frequent shared CDR3 nucleotide sequences of the TCR α and β chain between circulating CD8α-CD4 + SLAMF7 + T EM cells and pancreatic tissue of Patient #1. The frequency of clones expanded at baseline decreases after glucocorticoid treatment together with clinical improvement. Gray panels indicate missing nucleotide sequences. ( C ) Percentage decrease of circulating CD4 + T EM cells, total SLAMF7 + CD4 + CTLs, CD8α low CD4 + SLAMF7 + CTLs, and CD8α-CD4 + SLAMF7 + CTLs after six months of glucocorticoids (normalized to pre-treatment levels) is plotted against the IgG4-RD RI.
    Figure Legend Snippet: Clinical improvement induced by glucocorticoid treatment correlates with contraction of clonally expanded circulating CD8α − CD4 + SLAMF7 + T EM cells ( A ) Retrieval frequency of CDR3 amino acid sequences of TCRα and β chain from circulating CD8α-CD4 + SLAMF7 + T EM cells before and after treatment and in the pancreas of Patient #1, and in the blood, lacrimal gland, and lung tissues of Patient #5 show an oligoclonal pattern on Next-Generation Sequencing analysis. Circulating CD4 + T EM cells from a healthy donor are polyclonal. Red bars indicate the most frequently encountered sequences; violet bars represent the 10th most frequently encountered sequences; grey bars represent overall remaining CDR3 sequences. The higher the grey bar, the more polyclonal is the sample. Patient #1 also shows an increased polyclonality of the TCRα and β chain repertoire after treatment. ( B ) Ten most frequent shared CDR3 nucleotide sequences of the TCR α and β chain between circulating CD8α-CD4 + SLAMF7 + T EM cells and pancreatic tissue of Patient #1. The frequency of clones expanded at baseline decreases after glucocorticoid treatment together with clinical improvement. Gray panels indicate missing nucleotide sequences. ( C ) Percentage decrease of circulating CD4 + T EM cells, total SLAMF7 + CD4 + CTLs, CD8α low CD4 + SLAMF7 + CTLs, and CD8α-CD4 + SLAMF7 + CTLs after six months of glucocorticoids (normalized to pre-treatment levels) is plotted against the IgG4-RD RI.

    Techniques Used: Next-Generation Sequencing, Clone Assay

    28) Product Images from "Identification of Variants in Primary and Recurrent Glioblastoma Using a Cancer-Specific Gene Panel and Whole Exome Sequencing"

    Article Title: Identification of Variants in Primary and Recurrent Glioblastoma Using a Cancer-Specific Gene Panel and Whole Exome Sequencing

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0124178

    Overview of patient samples and sequencing methods. Sequencing for the AmpliSeq CCP samples was done using the Ion 318 Chip and whole exome sequencing was preformed using the Genome Analyzer II or HiSeq 2000, both from Illumina.
    Figure Legend Snippet: Overview of patient samples and sequencing methods. Sequencing for the AmpliSeq CCP samples was done using the Ion 318 Chip and whole exome sequencing was preformed using the Genome Analyzer II or HiSeq 2000, both from Illumina.

    Techniques Used: Sequencing, Chromatin Immunoprecipitation

    29) Product Images from "Comparative analysis of cytokine transcript profiles within mediastinal lymph node compartments of pigs after infection with porcine reproductive and respiratory syndrome genotype 1 strains differing in pathogenicity"

    Article Title: Comparative analysis of cytokine transcript profiles within mediastinal lymph node compartments of pigs after infection with porcine reproductive and respiratory syndrome genotype 1 strains differing in pathogenicity

    Journal: Veterinary Research

    doi: 10.1186/s13567-015-0161-8

    PRRSV viral load in mediastinal lymph node. PRRSV RNA was quantified by RT-qPCR and data represented by changes in the cycle threshold (Ct) in F and IF areas of Med-LN of LV (A) , 215–06 (B) and SU1-Bel (C) infected animals. Differences between distinct PRRSV-1 infected groups are showed for 3 (D) , 7 (E) , and 35 (F) days post-infection. This figure showed the median of each group ± SD. In control animals the viral RNA were not detected (data not shown). Identical superscript letters indicate no significant difference ( p > 0.05), whereas different superscript letters indicate statistically significant differences ( p
    Figure Legend Snippet: PRRSV viral load in mediastinal lymph node. PRRSV RNA was quantified by RT-qPCR and data represented by changes in the cycle threshold (Ct) in F and IF areas of Med-LN of LV (A) , 215–06 (B) and SU1-Bel (C) infected animals. Differences between distinct PRRSV-1 infected groups are showed for 3 (D) , 7 (E) , and 35 (F) days post-infection. This figure showed the median of each group ± SD. In control animals the viral RNA were not detected (data not shown). Identical superscript letters indicate no significant difference ( p > 0.05), whereas different superscript letters indicate statistically significant differences ( p

    Techniques Used: Quantitative RT-PCR, Infection

    30) Product Images from "A LAMP assay for the rapid and robust assessment of Wolbachia infection in Aedes aegypti under field and laboratory conditions"

    Article Title: A LAMP assay for the rapid and robust assessment of Wolbachia infection in Aedes aegypti under field and laboratory conditions

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0225321

    Concentration curves for (A) Ae . aegypti ITS1 (B) w AlbB wsp LAMP 5-primer sets (i.e. the forward loop primer of each has been removed). The horizontal axis shows the natural log of overall extracted DNA in the samples, while the vertical axis shows the peak amplification time (T p ) for each primer set under fresh reagent conditions. Linear regression lines are shown with 95% confidence intervals shaded.
    Figure Legend Snippet: Concentration curves for (A) Ae . aegypti ITS1 (B) w AlbB wsp LAMP 5-primer sets (i.e. the forward loop primer of each has been removed). The horizontal axis shows the natural log of overall extracted DNA in the samples, while the vertical axis shows the peak amplification time (T p ) for each primer set under fresh reagent conditions. Linear regression lines are shown with 95% confidence intervals shaded.

    Techniques Used: Concentration Assay, Amplification

    31) Product Images from "Quantification of Plasmid Copy Number with Single Colour Droplet Digital PCR"

    Article Title: Quantification of Plasmid Copy Number with Single Colour Droplet Digital PCR

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0169846

    Construction of the standard curves for bla and dxs . (A) The standard curves were calculated with serial 10-fold dilutions of pGEM-dxs, ranging from 1 × 10 5 to 1 × 10 9 copies μl -1 . Each standard dilution was amplified by qPCR using bla and dxs primer sets ( n = 2). For each gene, the determined C T values were plotted against the logarithm of their known initial copy number. A standard curve was generated by linear regression through these points. (B) Validation of the ΔΔC T calculation. The ΔC T deviation of bla vs. dxs was calculated for each dilution and plotted ( n = 2). Average ΔC T = average ± SD ( n = 10).
    Figure Legend Snippet: Construction of the standard curves for bla and dxs . (A) The standard curves were calculated with serial 10-fold dilutions of pGEM-dxs, ranging from 1 × 10 5 to 1 × 10 9 copies μl -1 . Each standard dilution was amplified by qPCR using bla and dxs primer sets ( n = 2). For each gene, the determined C T values were plotted against the logarithm of their known initial copy number. A standard curve was generated by linear regression through these points. (B) Validation of the ΔΔC T calculation. The ΔC T deviation of bla vs. dxs was calculated for each dilution and plotted ( n = 2). Average ΔC T = average ± SD ( n = 10).

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

    32) Product Images from "Glial cell type-specific changes in spinal dipeptidyl peptidase 4 expression and effects of its inhibitors in inflammatory and neuropatic pain"

    Article Title: Glial cell type-specific changes in spinal dipeptidyl peptidase 4 expression and effects of its inhibitors in inflammatory and neuropatic pain

    Journal: Scientific Reports

    doi: 10.1038/s41598-018-21799-8

    DPP4 immunoreactivity in glial cells in control, inflamed and neuropathic animals. Representative confocal images of the spinal dorsal horn obtained from control ( a ), inflamed ( b ) and neuropathic ( c ) animals demonstrate that DPP4 is expressed predominantly by glial cells (arrows: microglia, arrowheads: astrocytes). All the images are single optical sections. Scalebar: 10 μm. Integrated density values ( d ) demonstrate that the majority of DPP4-immunopositivity is related to GFAP-labelled astrocytes. A proportion of DPP4 labelling belongs to microgila and very few DPP4 is expressed by neuronal cell bodies. Inflammation significantly increased the DPP4 expression on astocytes but not on the other cell types. In contrast to inflammation DPP4 density was significantly higher on microglia and lower in neurons during neuropathy (Kruskal-Wallis One way ANOVA on ranks, Dunn’s Method, P
    Figure Legend Snippet: DPP4 immunoreactivity in glial cells in control, inflamed and neuropathic animals. Representative confocal images of the spinal dorsal horn obtained from control ( a ), inflamed ( b ) and neuropathic ( c ) animals demonstrate that DPP4 is expressed predominantly by glial cells (arrows: microglia, arrowheads: astrocytes). All the images are single optical sections. Scalebar: 10 μm. Integrated density values ( d ) demonstrate that the majority of DPP4-immunopositivity is related to GFAP-labelled astrocytes. A proportion of DPP4 labelling belongs to microgila and very few DPP4 is expressed by neuronal cell bodies. Inflammation significantly increased the DPP4 expression on astocytes but not on the other cell types. In contrast to inflammation DPP4 density was significantly higher on microglia and lower in neurons during neuropathy (Kruskal-Wallis One way ANOVA on ranks, Dunn’s Method, P

    Techniques Used: Expressing

    DPP4 immunoreactivity of different cell types in the spinal dorsal horn. DPP4-immunoreactive puncta appeared in Nissl stained neurons (row a), GFAP labelled astrocytes (row b) and IBA1 positive microglial cells (row c). Co-staining with mouse monoclonal antibody against the full length rat CD26 protein (DPP4mo, column 2) and polyclonal goat DPP4 antibody that was raised against the synthetic peptide C-PPHFDKSKKYP representing the internal region of DPP4 (DPP4gt, column 3) labelled the same puncta in all three cell types demonstrating the specificity of the two antibodies (arrows). All the images are single optical sections. Scalebar: 5 μm.
    Figure Legend Snippet: DPP4 immunoreactivity of different cell types in the spinal dorsal horn. DPP4-immunoreactive puncta appeared in Nissl stained neurons (row a), GFAP labelled astrocytes (row b) and IBA1 positive microglial cells (row c). Co-staining with mouse monoclonal antibody against the full length rat CD26 protein (DPP4mo, column 2) and polyclonal goat DPP4 antibody that was raised against the synthetic peptide C-PPHFDKSKKYP representing the internal region of DPP4 (DPP4gt, column 3) labelled the same puncta in all three cell types demonstrating the specificity of the two antibodies (arrows). All the images are single optical sections. Scalebar: 5 μm.

    Techniques Used: Staining

    DPP4 mRNA and protein expression in the dorsal horn of control, carrageenan treated and neuropathic rats. DPP4 mRNA expression in the dorsal horn of the spinal cord assessed by qPCR ( a ) and in situ hybridization ( b ) did not show significant difference among the three experimental groups (mean ± SEM, n = 6–9, one-way ANOVA, P = 0.30 and P = 0.21 for qPCR and ISH, respectively). In Western-blot experiments goat DPP4 antibody labelled lane at 110 kDa in spinal dorsal horn lysates taken from naive, inflamed and neuropathic animals ( c . Significantly increased DPP4 protein levels were detected in carrageenan-induced inflammation measured both by Western-blotting ( d ) and quantitative immunohistochemistry ( e ). (Values are given as mean ± SEM, n = 7–10, one-way ANOVA followed by Holm-Sidac post hoc test: P = 0.023 for Western blot experiments and one-way ANOVA with Student-Neuman-Keuls post hoc test: P = 0.016 for densitometry).
    Figure Legend Snippet: DPP4 mRNA and protein expression in the dorsal horn of control, carrageenan treated and neuropathic rats. DPP4 mRNA expression in the dorsal horn of the spinal cord assessed by qPCR ( a ) and in situ hybridization ( b ) did not show significant difference among the three experimental groups (mean ± SEM, n = 6–9, one-way ANOVA, P = 0.30 and P = 0.21 for qPCR and ISH, respectively). In Western-blot experiments goat DPP4 antibody labelled lane at 110 kDa in spinal dorsal horn lysates taken from naive, inflamed and neuropathic animals ( c . Significantly increased DPP4 protein levels were detected in carrageenan-induced inflammation measured both by Western-blotting ( d ) and quantitative immunohistochemistry ( e ). (Values are given as mean ± SEM, n = 7–10, one-way ANOVA followed by Holm-Sidac post hoc test: P = 0.023 for Western blot experiments and one-way ANOVA with Student-Neuman-Keuls post hoc test: P = 0.016 for densitometry).

    Techniques Used: Expressing, Real-time Polymerase Chain Reaction, In Situ Hybridization, Western Blot, Immunohistochemistry

    Antihyperalgesic effect of DPP4 inhibitor IPI and vildagliptin in carrageenan-induced inflammation. ( a ) Antihyperalgesic effect of IPI has been completely abolished by co-administartion of CTAP, but ( b ) neither TIPP[Ψ] nor ( c ) gNTI altered IPI-evoked antihyperalgesia. ( d ) Antihyperalgesic effect of vildagliptin was significantly reduced by CTAP and ( f ) gNTI, but was completely eliminated when ( e ) TIPP[Ψ] was co-injected. Inhibitory effects of opioid antagonists on IPI and vildagliptin related antihyperalgesia are summarized on bar graphs ( g ) and ( h ) constructed from data recorded 210 min after i.t. drug application. Comparisons were made with two-way ANOVA, Bonferoni post hoc test; + p
    Figure Legend Snippet: Antihyperalgesic effect of DPP4 inhibitor IPI and vildagliptin in carrageenan-induced inflammation. ( a ) Antihyperalgesic effect of IPI has been completely abolished by co-administartion of CTAP, but ( b ) neither TIPP[Ψ] nor ( c ) gNTI altered IPI-evoked antihyperalgesia. ( d ) Antihyperalgesic effect of vildagliptin was significantly reduced by CTAP and ( f ) gNTI, but was completely eliminated when ( e ) TIPP[Ψ] was co-injected. Inhibitory effects of opioid antagonists on IPI and vildagliptin related antihyperalgesia are summarized on bar graphs ( g ) and ( h ) constructed from data recorded 210 min after i.t. drug application. Comparisons were made with two-way ANOVA, Bonferoni post hoc test; + p

    Techniques Used: Injection, Construct

    33) Product Images from "Differential Responses of Pattern Recognition Receptors to Outer Membrane Vesicles of Three Periodontal Pathogens"

    Article Title: Differential Responses of Pattern Recognition Receptors to Outer Membrane Vesicles of Three Periodontal Pathogens

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0151967

    TLR2 and TLR4 Activation by P . gingivalis , T . denticola and T . forsythia OMVs. HEK-Blue TLR2 (A, B, C) and TLR4 (D, E, F) Cell lines were incubated with 20μL of either OMVs or ligands Pam3CSK4 (10 μg/mL in 5 fold dilutions) and LPS-EB (10 μg/mL in 10 fold dilutions) respectively. Alkaline phosphatase activity was determined after 20 hours incubation at 620 nm on a spectrophotometer. Crude OMV (B, E) and purified OMV preparations standardised by OMV count (A, D) and general protein (C, F) were used to ascertain the effects of purification and standardisation methods on OMV immunogenicity. Lipoproteins were observed by GC-MS fatty acid analysis to determine the % fatty acid chain lengths present (G). OMVs were subjected to SDS-PAGE and lipopolysaccharide and lipooligosaccharide detected by Pro-Q Emerald 300 LPS Gel Stain (H). Glycoproteins were identified using SyproRuby protein stain (I). Purified P . gingivalis LPS was used as a positive control. Molecular mass markers (Novex SeeBlue Plus2 Prestained Standard) are indicated in kDa next to each sample.
    Figure Legend Snippet: TLR2 and TLR4 Activation by P . gingivalis , T . denticola and T . forsythia OMVs. HEK-Blue TLR2 (A, B, C) and TLR4 (D, E, F) Cell lines were incubated with 20μL of either OMVs or ligands Pam3CSK4 (10 μg/mL in 5 fold dilutions) and LPS-EB (10 μg/mL in 10 fold dilutions) respectively. Alkaline phosphatase activity was determined after 20 hours incubation at 620 nm on a spectrophotometer. Crude OMV (B, E) and purified OMV preparations standardised by OMV count (A, D) and general protein (C, F) were used to ascertain the effects of purification and standardisation methods on OMV immunogenicity. Lipoproteins were observed by GC-MS fatty acid analysis to determine the % fatty acid chain lengths present (G). OMVs were subjected to SDS-PAGE and lipopolysaccharide and lipooligosaccharide detected by Pro-Q Emerald 300 LPS Gel Stain (H). Glycoproteins were identified using SyproRuby protein stain (I). Purified P . gingivalis LPS was used as a positive control. Molecular mass markers (Novex SeeBlue Plus2 Prestained Standard) are indicated in kDa next to each sample.

    Techniques Used: Activation Assay, Incubation, Activity Assay, Spectrophotometry, Purification, Gas Chromatography-Mass Spectrometry, SDS Page, Staining, Positive Control

    Isolation of highly purified P . gingivalis , T . denticola and T . forsythia OMVs. OptiPrep density gradient centrifugation of a crude T . forsythia OMV preparation (A1) separated non-OMV associated material (bottom of density gradient) from highly purified OMVs isolated at a higher density. A blank OptiPrep density gradient is shown in A2. SDS-PAGE of P . gingivalis (B), T . denticola (E F) and T . forsythia (I J) Optiprep density gradient fractions taken in eight 1.5 mL gradient fractions from top to bottom following centrifugation. Gradient fractions were subjected to SDS-PAGE and protein bands visualized by SimplyBlue or SyproRuby staining. Molecular mass markers (Novex SeeBlue Plus2 Prestained Standard) are indicated in kDa. OMVs were observed using transmission electron microscopy (TEM) (C G K). Vesicle size and range was further confirmed using dynamic light scattering (DLS) on pooled and washed OMV containing gradient fractions (D H L).
    Figure Legend Snippet: Isolation of highly purified P . gingivalis , T . denticola and T . forsythia OMVs. OptiPrep density gradient centrifugation of a crude T . forsythia OMV preparation (A1) separated non-OMV associated material (bottom of density gradient) from highly purified OMVs isolated at a higher density. A blank OptiPrep density gradient is shown in A2. SDS-PAGE of P . gingivalis (B), T . denticola (E F) and T . forsythia (I J) Optiprep density gradient fractions taken in eight 1.5 mL gradient fractions from top to bottom following centrifugation. Gradient fractions were subjected to SDS-PAGE and protein bands visualized by SimplyBlue or SyproRuby staining. Molecular mass markers (Novex SeeBlue Plus2 Prestained Standard) are indicated in kDa. OMVs were observed using transmission electron microscopy (TEM) (C G K). Vesicle size and range was further confirmed using dynamic light scattering (DLS) on pooled and washed OMV containing gradient fractions (D H L).

    Techniques Used: Isolation, Purification, Gradient Centrifugation, SDS Page, Centrifugation, Staining, Transmission Assay, Electron Microscopy, Transmission Electron Microscopy

    34) Product Images from "Circulating microRNA Associated to Different Stages of Liver Steatosis in Prader–Willi Syndrome and Non-Syndromic Obesity"

    Article Title: Circulating microRNA Associated to Different Stages of Liver Steatosis in Prader–Willi Syndrome and Non-Syndromic Obesity

    Journal: Journal of Clinical Medicine

    doi: 10.3390/jcm9041123

    ROC curve analysis of serum miRNAs discriminating PWS+ grade 1 and PWS+ grade 2/3. ( A ) ROC curves for miR-93-5p (60% sensitivity and 52% specificity) and miR-106b-5p (70% sensitivity and 68% specificity). ( B ) ROC curves illustrating the performance of the miRNA panel in discriminating grade 1 from grade 2/3 steatosis in PWS+. PWS+, Prader–Willi Syndrome with steatosis AUC, area under the ROC curve; CI, confidence interval.
    Figure Legend Snippet: ROC curve analysis of serum miRNAs discriminating PWS+ grade 1 and PWS+ grade 2/3. ( A ) ROC curves for miR-93-5p (60% sensitivity and 52% specificity) and miR-106b-5p (70% sensitivity and 68% specificity). ( B ) ROC curves illustrating the performance of the miRNA panel in discriminating grade 1 from grade 2/3 steatosis in PWS+. PWS+, Prader–Willi Syndrome with steatosis AUC, area under the ROC curve; CI, confidence interval.

    Techniques Used:

    Receiver operating characteristic (ROC) curve analysis for single microRNAs ( A ) and combinations (B: miR-122-5p, miR-151a, miR-92a-3p) to discriminate PWS from OB subjects. ( B ) The AUC for the combination of the three miRNA was determined as 0.81 (0.64-0.90, 95% CI) with a sensitivity of 77.7% and a specificity of 71.4%, at a cut-off 0.56 (logit model formula: –1.34 + 2.97 × miR-122-5p - 0.57 × miR-92a_3p + 1.62 × miR-151a-5p). AUC, area under the ROC curve; CI, confidence interval.
    Figure Legend Snippet: Receiver operating characteristic (ROC) curve analysis for single microRNAs ( A ) and combinations (B: miR-122-5p, miR-151a, miR-92a-3p) to discriminate PWS from OB subjects. ( B ) The AUC for the combination of the three miRNA was determined as 0.81 (0.64-0.90, 95% CI) with a sensitivity of 77.7% and a specificity of 71.4%, at a cut-off 0.56 (logit model formula: –1.34 + 2.97 × miR-122-5p - 0.57 × miR-92a_3p + 1.62 × miR-151a-5p). AUC, area under the ROC curve; CI, confidence interval.

    Techniques Used:

    ROC curve analysis of serum miRNAs discriminating grade 1 PWS+ from OBS+. ( A ) ROC curves for the single miRNA candidates, miR-151a, miR-92a-3p, miR-106b-5p, and miR-93-5p with optimal sensitivity/specificity value of 80/85%, 80/78%, 80/71%, and 70/78%, respectively. ( B ) The AUC for the miRNA panel was 0.82 (95% CI (0.50-0.94), cut-offs 0.82, with a sensitivity of 85.7%, and a specificity of 80% (logit model formula: –2.34 + 2.48 * miR-92a-3p + 1.47 × miR-151a-5p + 1.97 × miR-106b-5p - 2.55 × miR-93-5p. OBS+, obese subjects with steatosis; PWS+, Prader–Willi Syndrome with steatosis AUC, area under the ROC curve; CI, confidence interval.
    Figure Legend Snippet: ROC curve analysis of serum miRNAs discriminating grade 1 PWS+ from OBS+. ( A ) ROC curves for the single miRNA candidates, miR-151a, miR-92a-3p, miR-106b-5p, and miR-93-5p with optimal sensitivity/specificity value of 80/85%, 80/78%, 80/71%, and 70/78%, respectively. ( B ) The AUC for the miRNA panel was 0.82 (95% CI (0.50-0.94), cut-offs 0.82, with a sensitivity of 85.7%, and a specificity of 80% (logit model formula: –2.34 + 2.48 * miR-92a-3p + 1.47 × miR-151a-5p + 1.97 × miR-106b-5p - 2.55 × miR-93-5p. OBS+, obese subjects with steatosis; PWS+, Prader–Willi Syndrome with steatosis AUC, area under the ROC curve; CI, confidence interval.

    Techniques Used:

    35) Product Images from "Epigenetic resetting of human pluripotency"

    Article Title: Epigenetic resetting of human pluripotency

    Journal: Development (Cambridge, England)

    doi: 10.1242/dev.146811

    Transcriptome analysis of reset PSCs. (A) Principal component analysis (PCA) of whole-transcriptome RNA-seq data from the indicated cell lines. (B) t-SNE analysis of RNA-seq data. (C) Heatmap of differentially expressed genes between chemically reset (cR) and embryo-derived HNES cells (naïve) compared with conventional hPSCs (primed). Genes unregulated in naïve cells are shown, ranked by log 2 fold-change (FC). Values displayed correspond to the average expression level in each sample group scaled by the mean expression of each gene. (D) Heatmap showing expression of all transposon families that are differentially expressed (log 2 FC > 1.5, P
    Figure Legend Snippet: Transcriptome analysis of reset PSCs. (A) Principal component analysis (PCA) of whole-transcriptome RNA-seq data from the indicated cell lines. (B) t-SNE analysis of RNA-seq data. (C) Heatmap of differentially expressed genes between chemically reset (cR) and embryo-derived HNES cells (naïve) compared with conventional hPSCs (primed). Genes unregulated in naïve cells are shown, ranked by log 2 fold-change (FC). Values displayed correspond to the average expression level in each sample group scaled by the mean expression of each gene. (D) Heatmap showing expression of all transposon families that are differentially expressed (log 2 FC > 1.5, P

    Techniques Used: RNA Sequencing Assay, Derivative Assay, Expressing

    36) Product Images from "Generation and Neuronal Differentiation of hiPSCs From Patients With Myotonic Dystrophy Type 2"

    Article Title: Generation and Neuronal Differentiation of hiPSCs From Patients With Myotonic Dystrophy Type 2

    Journal: Frontiers in Physiology

    doi: 10.3389/fphys.2018.00967

    Detection of DM2 mutation at DNA, expression of CNBP gene and RNA level during differentiation. (A) Percentages of nuclear foci (1–4 or > 5) in hiPSCs and NP showing the increase of foci number along their differentiation process. (B) LR-PCR followed by hybridization with a (CTG) 5 -radioactively labeled probe on DNA extracted from DM2 hiPSCs and NPs, arrows indicated CNBP normal and expanded alleles. (C) RT-qPCR assay for CNBP expression in DM2 hiPSCs and NPs. β-actin is used as reference gene.
    Figure Legend Snippet: Detection of DM2 mutation at DNA, expression of CNBP gene and RNA level during differentiation. (A) Percentages of nuclear foci (1–4 or > 5) in hiPSCs and NP showing the increase of foci number along their differentiation process. (B) LR-PCR followed by hybridization with a (CTG) 5 -radioactively labeled probe on DNA extracted from DM2 hiPSCs and NPs, arrows indicated CNBP normal and expanded alleles. (C) RT-qPCR assay for CNBP expression in DM2 hiPSCs and NPs. β-actin is used as reference gene.

    Techniques Used: Mutagenesis, Expressing, Polymerase Chain Reaction, Hybridization, CTG Assay, Labeling, Quantitative RT-PCR

    37) Product Images from "DNA extraction approaches substantially influence the assessment of the human breast milk microbiome"

    Article Title: DNA extraction approaches substantially influence the assessment of the human breast milk microbiome

    Journal: Scientific Reports

    doi: 10.1038/s41598-019-55568-y

    DNA concentration (ng/µL) of mock breast milk ( a ) and human breast milk samples ( b ) as determined by Qubit dsDNA HS Assay kit. The dashed line represents the expected DNA concentration of 5.49 ng/uL, based on the bacteria added to the mock breast milk. Expected concentration was calculated based on the size of each bacterial genome, the weight of a base pair, and the CFU of each bacteria. *P
    Figure Legend Snippet: DNA concentration (ng/µL) of mock breast milk ( a ) and human breast milk samples ( b ) as determined by Qubit dsDNA HS Assay kit. The dashed line represents the expected DNA concentration of 5.49 ng/uL, based on the bacteria added to the mock breast milk. Expected concentration was calculated based on the size of each bacterial genome, the weight of a base pair, and the CFU of each bacteria. *P

    Techniques Used: Concentration Assay

    38) Product Images from "RNA-sequence data normalization through in silico prediction of reference genes: the bacterial response to DNA damage as case study"

    Article Title: RNA-sequence data normalization through in silico prediction of reference genes: the bacterial response to DNA damage as case study

    Journal: BioData Mining

    doi: 10.1186/s13040-017-0150-8

    Experimental design and evaluation of moose 2 . a Triplicate E. coli MG1655 cultures were treated with MMC, and total RNA samples were sequenced on an Illumina platform. b Variation plot for expression of six established reference genes as measured by qRT-PCR. Ct, cycle threshold. c Correlation between six established reference genes over all non-normalized RNA-seq samples, sorted by transcript abundance. The lowly expressed cysG , idnT , and hcaT are positively correlated (shown in yellow), but negatively correlated with the more abundant ihfB and rrsA . The colour scale bar reflects Pearson’s rho. d Log 2 -ratio distribution in cross-condition comparisons, shown for RPKM, Upper Quartile (UQ), Trimmed-Mean of M-values (TMM), DESeq2, and moose 2 . e Hierarchical clustering based on the Euclidean distance between RNA-seq samples. Matched conditions indicated as blue (0 min), black (30 min), and red (90 min)
    Figure Legend Snippet: Experimental design and evaluation of moose 2 . a Triplicate E. coli MG1655 cultures were treated with MMC, and total RNA samples were sequenced on an Illumina platform. b Variation plot for expression of six established reference genes as measured by qRT-PCR. Ct, cycle threshold. c Correlation between six established reference genes over all non-normalized RNA-seq samples, sorted by transcript abundance. The lowly expressed cysG , idnT , and hcaT are positively correlated (shown in yellow), but negatively correlated with the more abundant ihfB and rrsA . The colour scale bar reflects Pearson’s rho. d Log 2 -ratio distribution in cross-condition comparisons, shown for RPKM, Upper Quartile (UQ), Trimmed-Mean of M-values (TMM), DESeq2, and moose 2 . e Hierarchical clustering based on the Euclidean distance between RNA-seq samples. Matched conditions indicated as blue (0 min), black (30 min), and red (90 min)

    Techniques Used: Expressing, Quantitative RT-PCR, RNA Sequencing Assay

    39) Product Images from "Comparison of Pre-Analytical FFPE Sample Preparation Methods and Their Impact on Massively Parallel Sequencing in Routine Diagnostics"

    Article Title: Comparison of Pre-Analytical FFPE Sample Preparation Methods and Their Impact on Massively Parallel Sequencing in Routine Diagnostics

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0104566

    Comparison of five automated DNA extraction systems. Illustration of relative DNA concentrations of samples 1–10 measured by the NanoDrop 2000c spectrophotometer (A) and Qubit 2.0 fluorometer (B). The concentrations of the Maxwell 16 extracts were set to 100% for each sample.
    Figure Legend Snippet: Comparison of five automated DNA extraction systems. Illustration of relative DNA concentrations of samples 1–10 measured by the NanoDrop 2000c spectrophotometer (A) and Qubit 2.0 fluorometer (B). The concentrations of the Maxwell 16 extracts were set to 100% for each sample.

    Techniques Used: DNA Extraction, Spectrophotometry

    40) Product Images from "Comparison of UV spectrometry and fluorometry-based methods for quantification of cell-free DNA in red cell components"

    Article Title: Comparison of UV spectrometry and fluorometry-based methods for quantification of cell-free DNA in red cell components

    Journal: Asian Journal of Transfusion Science

    doi: 10.4103/ajts.AJTS_90_19

    Scatter plots showing comparison of cell-free DNA estimation by NanoDrop and Qubit 2.0NanoDrop and Qubit 2.0
    Figure Legend Snippet: Scatter plots showing comparison of cell-free DNA estimation by NanoDrop and Qubit 2.0NanoDrop and Qubit 2.0

    Techniques Used:

    Related Articles

    Fluorescence:

    Article Title: Validating DNA Extraction Protocols for Bentonite Clay
    Article Snippet: .. Genomic DNA concentration was determined using the Qubit dsDNA High Sensitivity (HS) assay kit (Invitrogen, Carlsbad, CA, USA) with fluorescence measured on a FilterMax F5 MultiMode plate reader (Molecular Devices, San Jose, CA, USA) at excitation and emission wavelengths of 485 and 525 nm, respectively. .. The quality of extracted E. coli DNA was assessed using a NanoDrop 2000 spectrophotometer (Thermo Scientific, Waltham, MA, USA) and pulsed-field gel electrophoresis (PFGE) using a CHEF Mapper XA apparatus (Bio-Rad, Hercules, CA, USA).

    Sensitive Assay:

    Article Title: Enhanced and controlled chromatin extraction from FFPE tissues and the application to ChIP-seq
    Article Snippet: .. DNAs were purified using Min-Elute PCR purification kit after the treatment of RNase A and proteinase K. DNA amount was measured by the Qubit dsDNA High Sensitivity assay (Invitrogen, Q32851), and DNA size was analyzed by the Fragment Analyzer (Advanced Analytical Technologies; AATI; Ankeny, IA) using the High Sensitivity NGS Fragment Analysis Kit (Cat. #DNF-486). .. For the comparison of soluble chromatin yield, the ChIP-IT® FFPE Chromatin Preparation Kit (Active Motif, Cat # 53030) was utilized according to the manufacture’s instructions.

    Article Title: Maintaining Breast Cancer Specimen Integrity and Individual or Simultaneous Extraction of Quality DNA, RNA, and Proteins from Allprotect-Stabilized and Nonstabilized Tissue Samples
    Article Snippet: .. DNA was quantified using Qubit [also known as Quant-iT™ dsDNA HS (High Sensitivity) Assay Kit; Invitrogen], Nanodrop 2000™, Thermo Scientific (designated Nanodrop1), and NanoDrop ND-1000™, NanoDrop Technologies (designated Nanodrop2), as per manufacturer's instructions. .. Both the ratio of absorbance at 260 and 280 nm (A260/280 ) and gel electrophoresis were utilized to ascertain the quality and purity of DNA isolates.

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application
    Article Snippet: .. After treatment with RNase A and proteinase K, DNA was purified by Qiagen MinElute PCR Purification Kit (Cat. # 28006, Valencia, CA) and quantified using Qubit dsDNA High Sensitivity assay (Invitrogen, Q32851). .. To check the size of input chromatin, purified input DNA was analyzed by Fragment Analyzer (Advanced Analytical Technologies; AATI; Ankeny, IA) using the High Sensitivity NGS Fragment Analysis Kit (Cat. # DNF-486).

    Polymerase Chain Reaction:

    Article Title: Enhanced and controlled chromatin extraction from FFPE tissues and the application to ChIP-seq
    Article Snippet: .. DNAs were purified using Min-Elute PCR purification kit after the treatment of RNase A and proteinase K. DNA amount was measured by the Qubit dsDNA High Sensitivity assay (Invitrogen, Q32851), and DNA size was analyzed by the Fragment Analyzer (Advanced Analytical Technologies; AATI; Ankeny, IA) using the High Sensitivity NGS Fragment Analysis Kit (Cat. #DNF-486). .. For the comparison of soluble chromatin yield, the ChIP-IT® FFPE Chromatin Preparation Kit (Active Motif, Cat # 53030) was utilized according to the manufacture’s instructions.

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application
    Article Snippet: .. After treatment with RNase A and proteinase K, DNA was purified by Qiagen MinElute PCR Purification Kit (Cat. # 28006, Valencia, CA) and quantified using Qubit dsDNA High Sensitivity assay (Invitrogen, Q32851). .. To check the size of input chromatin, purified input DNA was analyzed by Fragment Analyzer (Advanced Analytical Technologies; AATI; Ankeny, IA) using the High Sensitivity NGS Fragment Analysis Kit (Cat. # DNF-486).

    Next-Generation Sequencing:

    Article Title: Enhanced and controlled chromatin extraction from FFPE tissues and the application to ChIP-seq
    Article Snippet: .. DNAs were purified using Min-Elute PCR purification kit after the treatment of RNase A and proteinase K. DNA amount was measured by the Qubit dsDNA High Sensitivity assay (Invitrogen, Q32851), and DNA size was analyzed by the Fragment Analyzer (Advanced Analytical Technologies; AATI; Ankeny, IA) using the High Sensitivity NGS Fragment Analysis Kit (Cat. #DNF-486). .. For the comparison of soluble chromatin yield, the ChIP-IT® FFPE Chromatin Preparation Kit (Active Motif, Cat # 53030) was utilized according to the manufacture’s instructions.

    Purification:

    Article Title: Enhanced and controlled chromatin extraction from FFPE tissues and the application to ChIP-seq
    Article Snippet: .. DNAs were purified using Min-Elute PCR purification kit after the treatment of RNase A and proteinase K. DNA amount was measured by the Qubit dsDNA High Sensitivity assay (Invitrogen, Q32851), and DNA size was analyzed by the Fragment Analyzer (Advanced Analytical Technologies; AATI; Ankeny, IA) using the High Sensitivity NGS Fragment Analysis Kit (Cat. #DNF-486). .. For the comparison of soluble chromatin yield, the ChIP-IT® FFPE Chromatin Preparation Kit (Active Motif, Cat # 53030) was utilized according to the manufacture’s instructions.

    Article Title: Purification of nanogram-range immunoprecipitated DNA in ChIP-seq application
    Article Snippet: .. After treatment with RNase A and proteinase K, DNA was purified by Qiagen MinElute PCR Purification Kit (Cat. # 28006, Valencia, CA) and quantified using Qubit dsDNA High Sensitivity assay (Invitrogen, Q32851). .. To check the size of input chromatin, purified input DNA was analyzed by Fragment Analyzer (Advanced Analytical Technologies; AATI; Ankeny, IA) using the High Sensitivity NGS Fragment Analysis Kit (Cat. # DNF-486).

    Concentration Assay:

    Article Title: Validating DNA Extraction Protocols for Bentonite Clay
    Article Snippet: .. Genomic DNA concentration was determined using the Qubit dsDNA High Sensitivity (HS) assay kit (Invitrogen, Carlsbad, CA, USA) with fluorescence measured on a FilterMax F5 MultiMode plate reader (Molecular Devices, San Jose, CA, USA) at excitation and emission wavelengths of 485 and 525 nm, respectively. .. The quality of extracted E. coli DNA was assessed using a NanoDrop 2000 spectrophotometer (Thermo Scientific, Waltham, MA, USA) and pulsed-field gel electrophoresis (PFGE) using a CHEF Mapper XA apparatus (Bio-Rad, Hercules, CA, USA).

    Formalin-fixed Paraffin-Embedded:

    Article Title: Functional DNA quantification guides accurate next-generation sequencing mutation detection in formalin-fixed, paraffin-embedded tumor biopsies
    Article Snippet: .. DNA quantification using the Qubit® 2.0 Fluorometer A total of 165 FFPE samples were diluted to 5 ng/μl and quantified using the Qubit dsDNA HS Assay Kit (catalogue number Q32854, Life Technologies) and the Qubit 2.0 Fluorometer (Life Technologies) per the manufacturer’s instruction. .. Briefly, 1 μl of DNA sample at 5 ng/μl was diluted 200-fold in Qubit dsDNA HS buffer in clear plastic Qubit Assay Tubes (catalogue number Q32856, Life Technologies) and measured on the fluorometer.

    IA:

    Article Title: Enhanced and controlled chromatin extraction from FFPE tissues and the application to ChIP-seq
    Article Snippet: .. DNAs were purified using Min-Elute PCR purification kit after the treatment of RNase A and proteinase K. DNA amount was measured by the Qubit dsDNA High Sensitivity assay (Invitrogen, Q32851), and DNA size was analyzed by the Fragment Analyzer (Advanced Analytical Technologies; AATI; Ankeny, IA) using the High Sensitivity NGS Fragment Analysis Kit (Cat. #DNF-486). .. For the comparison of soluble chromatin yield, the ChIP-IT® FFPE Chromatin Preparation Kit (Active Motif, Cat # 53030) was utilized according to the manufacture’s instructions.

    other:

    Article Title: Biases during DNA extraction affect bacterial and archaeal community profile of anaerobic digestion samples
    Article Snippet: DNA yield was measured using two methods: Nanodrop (Nanodrop One, Thermoscientific, USA) and the Qubit fluorometer (Invitrogen, USA, using the Qubit® dsDNA HS Assay Kit).

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