platinum taq dna polymerase  (Thermo Fisher)


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    Structured Review

    Thermo Fisher platinum taq dna polymerase
    Workflow of reverse transcription and amplification of cfmRNA. cfmRNA, spiked-in with luciferase RNA control, was reverse transcript with Superscript™ III Reverse Transcriptase (Invitrogen, Cat no. 18080044). The product was amplified with rhPCR primers and Platinum™ <t>Taq</t> <t>DNA</t> Polymerase (Invitrogen, Cat no. 10966) using emulsion and CoT PCR. The residual primers were removed with Exonuclease I (New England Biolabs, Cat no. M0293). Amplified products were used for qPCR quantification and sequencing.
    Platinum Taq Dna Polymerase, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 39 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Images

    1) Product Images from "Non-Invasive Characterization of the Pancreas During Bariatric Surgery via Circulating Pancreatic Specific Cell-free Messenger RNA"

    Article Title: Non-Invasive Characterization of the Pancreas During Bariatric Surgery via Circulating Pancreatic Specific Cell-free Messenger RNA

    Journal: Frontiers in Genetics

    doi: 10.3389/fgene.2021.742496

    Workflow of reverse transcription and amplification of cfmRNA. cfmRNA, spiked-in with luciferase RNA control, was reverse transcript with Superscript™ III Reverse Transcriptase (Invitrogen, Cat no. 18080044). The product was amplified with rhPCR primers and Platinum™ Taq DNA Polymerase (Invitrogen, Cat no. 10966) using emulsion and CoT PCR. The residual primers were removed with Exonuclease I (New England Biolabs, Cat no. M0293). Amplified products were used for qPCR quantification and sequencing.
    Figure Legend Snippet: Workflow of reverse transcription and amplification of cfmRNA. cfmRNA, spiked-in with luciferase RNA control, was reverse transcript with Superscript™ III Reverse Transcriptase (Invitrogen, Cat no. 18080044). The product was amplified with rhPCR primers and Platinum™ Taq DNA Polymerase (Invitrogen, Cat no. 10966) using emulsion and CoT PCR. The residual primers were removed with Exonuclease I (New England Biolabs, Cat no. M0293). Amplified products were used for qPCR quantification and sequencing.

    Techniques Used: Amplification, Luciferase, RNase H-dependent PCR, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Sequencing

    2) Product Images from "Novel Algorithms Reveal Streptococcal Transcriptomes and Clues about Undefined Genes"

    Article Title: Novel Algorithms Reveal Streptococcal Transcriptomes and Clues about Undefined Genes

    Journal: PLoS Computational Biology

    doi: 10.1371/journal.pcbi.0030132

    Confirmation of spy0129 Deletion Mutant and Adherence Assay Top panel is a schematic representation of the spy0127–0130 cluster. Position of primers for RT-PCR are indicated by arrows. (A) Results of RT-PCR analysis on mRNA from two putative deletion mutants (Δ1 and Δ2) and the parental SF370 strain (P). mRNA was isolated from mid log-phase or stationary phase cells (indicated below panel) and reverse-transcribed with two primer combinations, which are indicated at the top of the lanes as primer set A (0129F–0129R) and primer set B (0129F–0130R). cDNA products were separated on a 1% agarose gel and visualized by ethidium bromide staining. The expected sizes of resulting cDNAs from SF370 using primer set A is 365 bp and using primer set B is 1200 bp. Control reactions (C) containing mid-log phase mRNA and Taq DNA polymerase instead of reverse transcriptase are indicated. Lane 1 contains 1 kb Plus DNA ladder (1 μg; Invitrogen). (B) Results of the pharyngeal cell adherence assay (detailed in Methods), comparing parental strain SF370 to the spy0129–0130 isogenic mutant, SF370Δ spy0129 .2 (abbreviated as Δ spy0129 ). Adherent streptococci are reported as the percentage of total number of streptococci added as inoculum to pharyngeal cell monolayers. Statistical significance (reported as p -value) was determined by Student's t- test.
    Figure Legend Snippet: Confirmation of spy0129 Deletion Mutant and Adherence Assay Top panel is a schematic representation of the spy0127–0130 cluster. Position of primers for RT-PCR are indicated by arrows. (A) Results of RT-PCR analysis on mRNA from two putative deletion mutants (Δ1 and Δ2) and the parental SF370 strain (P). mRNA was isolated from mid log-phase or stationary phase cells (indicated below panel) and reverse-transcribed with two primer combinations, which are indicated at the top of the lanes as primer set A (0129F–0129R) and primer set B (0129F–0130R). cDNA products were separated on a 1% agarose gel and visualized by ethidium bromide staining. The expected sizes of resulting cDNAs from SF370 using primer set A is 365 bp and using primer set B is 1200 bp. Control reactions (C) containing mid-log phase mRNA and Taq DNA polymerase instead of reverse transcriptase are indicated. Lane 1 contains 1 kb Plus DNA ladder (1 μg; Invitrogen). (B) Results of the pharyngeal cell adherence assay (detailed in Methods), comparing parental strain SF370 to the spy0129–0130 isogenic mutant, SF370Δ spy0129 .2 (abbreviated as Δ spy0129 ). Adherent streptococci are reported as the percentage of total number of streptococci added as inoculum to pharyngeal cell monolayers. Statistical significance (reported as p -value) was determined by Student's t- test.

    Techniques Used: Mutagenesis, Reverse Transcription Polymerase Chain Reaction, Isolation, Agarose Gel Electrophoresis, Staining

    Confirmation of speH Deletion Mutant and Pharyngeal Cell Adherence Assay (A) Results of RT-PCR and PCR analyses of total RNA preparations isolated from mid-log (OD = 0.4) and stationary phase (OD = 1) cultures of the ΔspeH deletion mutant (ΔL and ΔS, respectively), and stationary phase cultures of the SF370 parental strain (P). RNA was reverse-transcribed as described in Methods. To assess genomic DNA contamination, control reactions containing Taq DNA polymerase instead of reverse transcriptase were included. cDNA products were separated on a 1% agarose gel and visualized by ethidium bromide staining. Lanes containing products from either the RT-PCR or PCR analysis are designated at the bottom of the panel. Lanes labeled MW contain 1 kb Plus DNA ladder (1 μg; Invitrogen). (B) Results of the pharyngeal cell adherence assay (detailed in Methods), comparing parental strain SF370 with the deletion mutant SF370Δ speH (abbreviated Δ speH ). Adherent streptococci are reported as the percentage of total number of streptococci added as inoculum to pharyngeal cell monolayers. Statistical significance (reported as p value) was determined by Student's t- test.
    Figure Legend Snippet: Confirmation of speH Deletion Mutant and Pharyngeal Cell Adherence Assay (A) Results of RT-PCR and PCR analyses of total RNA preparations isolated from mid-log (OD = 0.4) and stationary phase (OD = 1) cultures of the ΔspeH deletion mutant (ΔL and ΔS, respectively), and stationary phase cultures of the SF370 parental strain (P). RNA was reverse-transcribed as described in Methods. To assess genomic DNA contamination, control reactions containing Taq DNA polymerase instead of reverse transcriptase were included. cDNA products were separated on a 1% agarose gel and visualized by ethidium bromide staining. Lanes containing products from either the RT-PCR or PCR analysis are designated at the bottom of the panel. Lanes labeled MW contain 1 kb Plus DNA ladder (1 μg; Invitrogen). (B) Results of the pharyngeal cell adherence assay (detailed in Methods), comparing parental strain SF370 with the deletion mutant SF370Δ speH (abbreviated Δ speH ). Adherent streptococci are reported as the percentage of total number of streptococci added as inoculum to pharyngeal cell monolayers. Statistical significance (reported as p value) was determined by Student's t- test.

    Techniques Used: Mutagenesis, Reverse Transcription Polymerase Chain Reaction, Polymerase Chain Reaction, Isolation, Agarose Gel Electrophoresis, Staining, Labeling

    3) Product Images from "Deletion of the CTRL2 Insulator in HSV-1 Results in the Decreased Expression of Genes Involved in Axonal Transport and Attenuates Reactivation In Vivo"

    Article Title: Deletion of the CTRL2 Insulator in HSV-1 Results in the Decreased Expression of Genes Involved in Axonal Transport and Attenuates Reactivation In Vivo

    Journal: Viruses

    doi: 10.3390/v14050909

    HSV-1 genome copies in rabbit ganglia in wt 17Syn+ and the ΔCTRL2 recombinant following infection. The number of HSV-1 genomes per ganglia was determined between wt 17Syn+ and ΔCTRL2 viral infections in rabbits. Relative copies of HSV-1 DNA polymerase were determined by PCR at 28 dpi. qPCR data is presented as a ratio of HSV-1 DNA pol/rabbit GAPDH. One-way ANOVA of independent samples was used to determine the statistical significance between the mutant and wt ( n = 6).
    Figure Legend Snippet: HSV-1 genome copies in rabbit ganglia in wt 17Syn+ and the ΔCTRL2 recombinant following infection. The number of HSV-1 genomes per ganglia was determined between wt 17Syn+ and ΔCTRL2 viral infections in rabbits. Relative copies of HSV-1 DNA polymerase were determined by PCR at 28 dpi. qPCR data is presented as a ratio of HSV-1 DNA pol/rabbit GAPDH. One-way ANOVA of independent samples was used to determine the statistical significance between the mutant and wt ( n = 6).

    Techniques Used: Recombinant, Infection, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Mutagenesis

    4) Product Images from "Simple allele-discriminating PCR for cost-effective and rapid genotyping and mapping"

    Article Title: Simple allele-discriminating PCR for cost-effective and rapid genotyping and mapping

    Journal: Plant Methods

    doi: 10.1186/1746-4811-5-1

    Adaptation of SAP for high throughput applications . (A) Diagram illustrating the Amplifluor SNP genotyping assay system. The allele-specific primer each has a unique 5' tail sequence that is identical to the 3' region of one of the Amplifluor SNP Universal Primers (FAM or JOE indicated by green and red arrows respectively). When combined with the common reverse primer, PCR amplification results in the synthesis of the tail sequence complement (thin red line). The Amplifluor ® SNPs Universal Primer then anneals specifically to the tail of reverse complement and is elongated by Taq Polymerase. Subsequent PCR cycles unfold the hairpin structure (indicated by filled circle) of the Amplifluor ® SNPs Universal Primers, which results in fluorescent signals. (B) A scatter plot showing results of a SAP-based Amplifluor SNP assay. X-axis represents the FAM signal measuring the amplification of WT LUG (+), and the Y-axis indicates the JOE signal that measures lug-16 -specific PCR amplification. Two types of controls were used. First, the manufacturer's template controls (GG, GT, TT) utilize FAM/JOE SNP primers and the control templates (GG, TT, and GT), both of which are provided by the manufacturer's kit. Second, the non-target control (NTC) uses water instead of DNA template. Results of three experimental samples ( lug-16/lug-16 , lug-16/+ , and +/+) are shown. DNA template was from known genotype. The experiment has been performed twice with similar results. The result from one such an experiment is shown.
    Figure Legend Snippet: Adaptation of SAP for high throughput applications . (A) Diagram illustrating the Amplifluor SNP genotyping assay system. The allele-specific primer each has a unique 5' tail sequence that is identical to the 3' region of one of the Amplifluor SNP Universal Primers (FAM or JOE indicated by green and red arrows respectively). When combined with the common reverse primer, PCR amplification results in the synthesis of the tail sequence complement (thin red line). The Amplifluor ® SNPs Universal Primer then anneals specifically to the tail of reverse complement and is elongated by Taq Polymerase. Subsequent PCR cycles unfold the hairpin structure (indicated by filled circle) of the Amplifluor ® SNPs Universal Primers, which results in fluorescent signals. (B) A scatter plot showing results of a SAP-based Amplifluor SNP assay. X-axis represents the FAM signal measuring the amplification of WT LUG (+), and the Y-axis indicates the JOE signal that measures lug-16 -specific PCR amplification. Two types of controls were used. First, the manufacturer's template controls (GG, GT, TT) utilize FAM/JOE SNP primers and the control templates (GG, TT, and GT), both of which are provided by the manufacturer's kit. Second, the non-target control (NTC) uses water instead of DNA template. Results of three experimental samples ( lug-16/lug-16 , lug-16/+ , and +/+) are shown. DNA template was from known genotype. The experiment has been performed twice with similar results. The result from one such an experiment is shown.

    Techniques Used: High Throughput Screening Assay, SNP Genotyping Assay, Sequencing, Polymerase Chain Reaction, Amplification

    5) Product Images from "Melt Analysis of Mismatch Amplification Mutation Assays (Melt-MAMA): A Functional Study of a Cost-Effective SNP Genotyping Assay in Bacterial Models"

    Article Title: Melt Analysis of Mismatch Amplification Mutation Assays (Melt-MAMA): A Functional Study of a Cost-Effective SNP Genotyping Assay in Bacterial Models

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0032866

    The principle of the Melt-MAMA PCR reaction. Four different scenarios involving two alternate SNP allele templates (I II vs. III IV) and the interaction of Allele-Specific (AS) PCR amplification using MAMA primers. The annealing of AS-MAMA primers to their allelic templates is shown with one primer labeled with a 5′ GC-clamp (Ia) whereas the other is not (IVa). (Ib and IVb) Taq Polymerase extends from the 3′ matched AS-MAMA primer despite the antepenultimate destabilizing nucleotide. (Ic and IVc) The second PCR cycle replicates from a newly synthesized DNA template made in the previous step (Ib and IVb). With the synthesized DNA serving as the template, a perfect primer-template complex is formed eliminating the antepenultimate destabilizing mismatch observed in Iab and IVab. At PCR endpoint (Id and IVd), the amplicons generated from the 3′ matched AS-MAMA primer greatly outnumbers the amplicons generated by the mismatched AS-MAMA primer. Temperature-dissociation curve plots (Ie and IVe) of each AS-PCR product (Iabcd, IIab and IIIab, IVabcd), showing the fluorescent intensity and the rate of fluorescent intensity change (derivative) as a function of temperature. For each allelic template reaction (I II vs. III IV), the melt profiles (Ie and IVe) show only a single change in fluorescent intensity. This indicates the amplification of the perfect-matched amplicon and little to no amplification of the mismatched amplicon. The GC –clamp “labeled” amplicons dissociate at higher temperatures (∼3°C to 5°C) than non-GC amplicons. Nonproductive primer annealing is shown for an AS-MAMA primer (IIa) and a GC-clamp AS-MAMA primer (IIIa) binding with their respective corresponding mismatched templates. The lack of Watson-Crick base pairing at two 3′ positions (the antepenultimate nucleotide at the 3′ end) of the AS primer introduces instability at this region (IIb and IIIb). This prevents efficient extension by the polymerase, which retards or prevents product amplification (Ie and IVe).
    Figure Legend Snippet: The principle of the Melt-MAMA PCR reaction. Four different scenarios involving two alternate SNP allele templates (I II vs. III IV) and the interaction of Allele-Specific (AS) PCR amplification using MAMA primers. The annealing of AS-MAMA primers to their allelic templates is shown with one primer labeled with a 5′ GC-clamp (Ia) whereas the other is not (IVa). (Ib and IVb) Taq Polymerase extends from the 3′ matched AS-MAMA primer despite the antepenultimate destabilizing nucleotide. (Ic and IVc) The second PCR cycle replicates from a newly synthesized DNA template made in the previous step (Ib and IVb). With the synthesized DNA serving as the template, a perfect primer-template complex is formed eliminating the antepenultimate destabilizing mismatch observed in Iab and IVab. At PCR endpoint (Id and IVd), the amplicons generated from the 3′ matched AS-MAMA primer greatly outnumbers the amplicons generated by the mismatched AS-MAMA primer. Temperature-dissociation curve plots (Ie and IVe) of each AS-PCR product (Iabcd, IIab and IIIab, IVabcd), showing the fluorescent intensity and the rate of fluorescent intensity change (derivative) as a function of temperature. For each allelic template reaction (I II vs. III IV), the melt profiles (Ie and IVe) show only a single change in fluorescent intensity. This indicates the amplification of the perfect-matched amplicon and little to no amplification of the mismatched amplicon. The GC –clamp “labeled” amplicons dissociate at higher temperatures (∼3°C to 5°C) than non-GC amplicons. Nonproductive primer annealing is shown for an AS-MAMA primer (IIa) and a GC-clamp AS-MAMA primer (IIIa) binding with their respective corresponding mismatched templates. The lack of Watson-Crick base pairing at two 3′ positions (the antepenultimate nucleotide at the 3′ end) of the AS primer introduces instability at this region (IIb and IIIb). This prevents efficient extension by the polymerase, which retards or prevents product amplification (Ie and IVe).

    Techniques Used: Polymerase Chain Reaction, Amplification, Labeling, Synthesized, Generated, Binding Assay

    6) Product Images from "Advancing uracil-excision based cloning towards an ideal technique for cloning PCR fragments"

    Article Title: Advancing uracil-excision based cloning towards an ideal technique for cloning PCR fragments

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkl635

    Characteristics and applications of the USER technique. ( a ) A comparison of the ability of DNA polymerases to amplify DNA fragments using uracil-free (-dU) and uracil-containing primers (+dU). Hm Taq: HotMaster™ Taq DNA Polymerase, Taq: Platinum ® Taq DNA Polymerase, Pwo: Pwo DNA Polymerase, Phusion: Phusion™ DNA Polymerase, PfuCx: PfuTurbo ® C x Hotstart DNA polymerase. ( b ) Functional expression of A.thaliana glucose transporter AtSTP1 in X.laevis oocytes from the Xenopus -specific USER compatible vector, pNB1u. H 2 O: control oocytes injected with water; STP1: oocytes injected with AtSTP1 cRNA. ( c ) Functional expression of the cyano fluorescence protein in leaf and root of A.thaliana from the CaMV 35S promoter using the USER compatible pCAMBIA230035Su vector (35S:CFP). Control: empty pCAMBIA230035Su.
    Figure Legend Snippet: Characteristics and applications of the USER technique. ( a ) A comparison of the ability of DNA polymerases to amplify DNA fragments using uracil-free (-dU) and uracil-containing primers (+dU). Hm Taq: HotMaster™ Taq DNA Polymerase, Taq: Platinum ® Taq DNA Polymerase, Pwo: Pwo DNA Polymerase, Phusion: Phusion™ DNA Polymerase, PfuCx: PfuTurbo ® C x Hotstart DNA polymerase. ( b ) Functional expression of A.thaliana glucose transporter AtSTP1 in X.laevis oocytes from the Xenopus -specific USER compatible vector, pNB1u. H 2 O: control oocytes injected with water; STP1: oocytes injected with AtSTP1 cRNA. ( c ) Functional expression of the cyano fluorescence protein in leaf and root of A.thaliana from the CaMV 35S promoter using the USER compatible pCAMBIA230035Su vector (35S:CFP). Control: empty pCAMBIA230035Su.

    Techniques Used: Functional Assay, Expressing, Plasmid Preparation, Injection, Fluorescence

    7) Product Images from "Cruzipain and Its Physiological Inhibitor, Chagasin, as a DNA-Based Therapeutic Vaccine Against Trypanosoma cruzi"

    Article Title: Cruzipain and Its Physiological Inhibitor, Chagasin, as a DNA-Based Therapeutic Vaccine Against Trypanosoma cruzi

    Journal: Frontiers in Immunology

    doi: 10.3389/fimmu.2020.565142

    Prevention of tissue damage by vaccine treatment during the acute phase of the parasite infection. (A) Enzymatic activity of CK and CK-MB enzymes represented as international units (IU/L). (B–E) Representative image of hematoxylin–eosin-stained skeletal muscle samples taken at day 100 after the last immunization; magnification: 40× and the inbox 100×. Animals showed (B) multiple confluent foci and necrosis with diffuse distribution (SControl); (C) nonconfluent mononuclear inflammatory infiltrate (SChg); (D) isolated foci of mononuclear inflammatory infiltrate (SCz); (E) few and isolated infiltrates (SChg+SCz). (F) Semi-quantified inflammation expressed as inflammation index: (1) isolated foci; (2) multiple nonconfluent foci; (3) inflammatory confluent foci; and (4) multiple diffuse foci ( 40 ). (G) Parasitism in skeletal and cardiac tissues by qPCR at day 100 after the last immunization. Parasite burden in each tissue was expressed as T. cruzi equivalents per 50 ng of total DNA referred to a calibration curve previously constructed containing known concentrations of T. cruzi epimastigotes. These results are representative of at least three independent experiments, each one being carried out with five animals per group. * p
    Figure Legend Snippet: Prevention of tissue damage by vaccine treatment during the acute phase of the parasite infection. (A) Enzymatic activity of CK and CK-MB enzymes represented as international units (IU/L). (B–E) Representative image of hematoxylin–eosin-stained skeletal muscle samples taken at day 100 after the last immunization; magnification: 40× and the inbox 100×. Animals showed (B) multiple confluent foci and necrosis with diffuse distribution (SControl); (C) nonconfluent mononuclear inflammatory infiltrate (SChg); (D) isolated foci of mononuclear inflammatory infiltrate (SCz); (E) few and isolated infiltrates (SChg+SCz). (F) Semi-quantified inflammation expressed as inflammation index: (1) isolated foci; (2) multiple nonconfluent foci; (3) inflammatory confluent foci; and (4) multiple diffuse foci ( 40 ). (G) Parasitism in skeletal and cardiac tissues by qPCR at day 100 after the last immunization. Parasite burden in each tissue was expressed as T. cruzi equivalents per 50 ng of total DNA referred to a calibration curve previously constructed containing known concentrations of T. cruzi epimastigotes. These results are representative of at least three independent experiments, each one being carried out with five animals per group. * p

    Techniques Used: Infection, Activity Assay, Staining, Isolation, Real-time Polymerase Chain Reaction, Construct

    Immune response in mice treated during the acute phase of T. cruzi infection. Mice infected with a sublethal dose of T. cruzi were treated at 0, 10, and 20 dpi with doses of Salmonella carrying the DNA of Chg and/or Cz together with the GM-CSF. Antibody immune response: Total IgG titers of (A) anti-rChg, (B) anti-rCz, and (C) anti-F105 were measured in sera of all groups obtained at 35 dpi. In vivo cellular immune response: Footpad thickness was measured before and 48 h after intradermal inoculation of (D) rChg or (E) rCz. The bars represent the average ( n = 5 per group) of three representative experiments. * p
    Figure Legend Snippet: Immune response in mice treated during the acute phase of T. cruzi infection. Mice infected with a sublethal dose of T. cruzi were treated at 0, 10, and 20 dpi with doses of Salmonella carrying the DNA of Chg and/or Cz together with the GM-CSF. Antibody immune response: Total IgG titers of (A) anti-rChg, (B) anti-rCz, and (C) anti-F105 were measured in sera of all groups obtained at 35 dpi. In vivo cellular immune response: Footpad thickness was measured before and 48 h after intradermal inoculation of (D) rChg or (E) rCz. The bars represent the average ( n = 5 per group) of three representative experiments. * p

    Techniques Used: Mouse Assay, Infection, In Vivo

    Prevention of T. cruzi -associated tissue damage by the vaccines administered at the chronic phase of the parasite infection. (A) Scheme of the chronic model of infection: C3H/HeN mice infected with T. cruzi received three doses of the SControl, SChg, SCz, or SChg+SCz on 100, 110, and 120 dpi ( n = 5 per group). (B) Enzymatic activity of CK and CK-MB enzymes represented as international units (IU/L). (C,D) Histopathological analysis of skeletal tissue samples at 220 dpi showed (C) confluent foci of mononuclear inflammatory infiltrate with necrosis of the muscle fibers in SControl; (D) isolated foci of mononuclear inflammatory cells with interstitial and perivascular predominance (SChg); (E) nonconfluent mononuclear cells surrounding muscle fibers and interstitial edema (SCz); and (F) isolated mononuclear cells at the interstitial and perivascular level (SChg+SCz). (G) Inflammation semi-quantified and expressed as an index of inflammation: (1) isolated foci; (2) multiple nonconfluent foci; (3) inflammatory confluent foci; and (4) multiple diffuse foci ( 40 ). (H) Parasite load in skeletal and cardiac tissues determined by qPCR at 220 dpi. Parasite burden in each tissue was expressed as T. cruzi equivalents per 50 ng of total DNA. Results were referred to a calibration curve previously constructed containing known concentrations of T. cruzi epimastigotes. These results are representative of at least three independent experiments. * p
    Figure Legend Snippet: Prevention of T. cruzi -associated tissue damage by the vaccines administered at the chronic phase of the parasite infection. (A) Scheme of the chronic model of infection: C3H/HeN mice infected with T. cruzi received three doses of the SControl, SChg, SCz, or SChg+SCz on 100, 110, and 120 dpi ( n = 5 per group). (B) Enzymatic activity of CK and CK-MB enzymes represented as international units (IU/L). (C,D) Histopathological analysis of skeletal tissue samples at 220 dpi showed (C) confluent foci of mononuclear inflammatory infiltrate with necrosis of the muscle fibers in SControl; (D) isolated foci of mononuclear inflammatory cells with interstitial and perivascular predominance (SChg); (E) nonconfluent mononuclear cells surrounding muscle fibers and interstitial edema (SCz); and (F) isolated mononuclear cells at the interstitial and perivascular level (SChg+SCz). (G) Inflammation semi-quantified and expressed as an index of inflammation: (1) isolated foci; (2) multiple nonconfluent foci; (3) inflammatory confluent foci; and (4) multiple diffuse foci ( 40 ). (H) Parasite load in skeletal and cardiac tissues determined by qPCR at 220 dpi. Parasite burden in each tissue was expressed as T. cruzi equivalents per 50 ng of total DNA. Results were referred to a calibration curve previously constructed containing known concentrations of T. cruzi epimastigotes. These results are representative of at least three independent experiments. * p

    Techniques Used: Infection, Mouse Assay, Activity Assay, Isolation, Real-time Polymerase Chain Reaction, Construct

    8) Product Images from "Comparative Methods to Improve the Detection of BRAF V600 Mutations in Highly Pigmented Melanoma Specimens"

    Article Title: Comparative Methods to Improve the Detection of BRAF V600 Mutations in Highly Pigmented Melanoma Specimens

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0158698

    Removal of the inhibitory effects of melanin on PCR amplification. (A) Increasing concentrations of synthetic melanin were added to DNA extracted from 1676 melanoma cell lines. (B) Increasing concentrations of BSA (ng/μl) were added to DNA extracted from cultured cells containing 40 or 80 ng/μl of melanin. (C) The effect of diluting DNA assessed in the presence of 40 ng/μl of melanin and either 1U or 2U of Taq polymerase. (D) NucleoSpin ® gDNA Clean-up XS Kit used on DNA extracted from cultured cells containing 40 or 80 ng/μl of melanin. PCR amplification of the DNA was monitored on 2% gel agarose electrophoresis with ethidium bromide staining. MW, molecular weight markers.
    Figure Legend Snippet: Removal of the inhibitory effects of melanin on PCR amplification. (A) Increasing concentrations of synthetic melanin were added to DNA extracted from 1676 melanoma cell lines. (B) Increasing concentrations of BSA (ng/μl) were added to DNA extracted from cultured cells containing 40 or 80 ng/μl of melanin. (C) The effect of diluting DNA assessed in the presence of 40 ng/μl of melanin and either 1U or 2U of Taq polymerase. (D) NucleoSpin ® gDNA Clean-up XS Kit used on DNA extracted from cultured cells containing 40 or 80 ng/μl of melanin. PCR amplification of the DNA was monitored on 2% gel agarose electrophoresis with ethidium bromide staining. MW, molecular weight markers.

    Techniques Used: Polymerase Chain Reaction, Amplification, Cell Culture, Electrophoresis, Staining, Molecular Weight

    9) Product Images from "A test of somatic mosaicism in the androgen receptor gene of Canada lynx (Lynx canadensis)"

    Article Title: A test of somatic mosaicism in the androgen receptor gene of Canada lynx (Lynx canadensis)

    Journal: BMC Genetics

    doi: 10.1186/s12863-015-0284-y

    Differential peak morphologies of androgen receptor alleles resulting from DNA dilution and reagent use. Lynx positive control DNA sample amplified with Invitrogen Taq DNA Polymerase and diluted to 1:10 ( a ), 1:20 ( b ), and 1:50 ( c ) ratios with deionized water. Lynx positive control DNA sample amplified with Invitrogen Platinum Taq DNA Polymerase (no dilution necessary) ( d )
    Figure Legend Snippet: Differential peak morphologies of androgen receptor alleles resulting from DNA dilution and reagent use. Lynx positive control DNA sample amplified with Invitrogen Taq DNA Polymerase and diluted to 1:10 ( a ), 1:20 ( b ), and 1:50 ( c ) ratios with deionized water. Lynx positive control DNA sample amplified with Invitrogen Platinum Taq DNA Polymerase (no dilution necessary) ( d )

    Techniques Used: Positive Control, Amplification

    10) Product Images from "Multiplex amplification enabled by selective circularization of large sets of genomic DNA fragments"

    Article Title: Multiplex amplification enabled by selective circularization of large sets of genomic DNA fragments

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gni070

    Selector construct and circularization reaction principles. ( A ) The selector consists of two oligonucleotides, one selector probe of ∼70 nt and one general vector oligonucleotide of 34 nt. The selector probe has two target-specific ends (black) of ∼18 nt complementary to the selected target and a primer-pair motif (gray) of 34 nt. The vector oligonucleotide is complementary to the primer-pair motif (gray). ( B ) The circularization procedure starts with digestion of the DNA to generate targets with defined ends. The digested DNA is then combined with selectors and the mixture is denaturated, allowing the selectors to hybridize to their respective targets. This can be performed in two different ways: (i) a selector probe hybridizes to both ends of the selected target and the ends are connected to the vector by ligation, or (ii) a selector probe hybridizes with one end to the 3′ end of the selected target and the other end to an internal sequence of the target, forming a branched structure that can be cleaved by the endonucleolytic activity of Taq DNA polymerase. Both ends of the selected single-stranded target are now ready for ligation to the vector oligonucleotide, forming a closed circular molecule. The circularization mixture is then exonuclease-treated and finally PCR-amplified using a common primer pair.
    Figure Legend Snippet: Selector construct and circularization reaction principles. ( A ) The selector consists of two oligonucleotides, one selector probe of ∼70 nt and one general vector oligonucleotide of 34 nt. The selector probe has two target-specific ends (black) of ∼18 nt complementary to the selected target and a primer-pair motif (gray) of 34 nt. The vector oligonucleotide is complementary to the primer-pair motif (gray). ( B ) The circularization procedure starts with digestion of the DNA to generate targets with defined ends. The digested DNA is then combined with selectors and the mixture is denaturated, allowing the selectors to hybridize to their respective targets. This can be performed in two different ways: (i) a selector probe hybridizes to both ends of the selected target and the ends are connected to the vector by ligation, or (ii) a selector probe hybridizes with one end to the 3′ end of the selected target and the other end to an internal sequence of the target, forming a branched structure that can be cleaved by the endonucleolytic activity of Taq DNA polymerase. Both ends of the selected single-stranded target are now ready for ligation to the vector oligonucleotide, forming a closed circular molecule. The circularization mixture is then exonuclease-treated and finally PCR-amplified using a common primer pair.

    Techniques Used: Construct, Plasmid Preparation, Ligation, Sequencing, Activity Assay, Polymerase Chain Reaction, Amplification

    11) Product Images from "PCR hot start using primers with the structure of molecular beacons (hairpin-like structure)"

    Article Title: PCR hot start using primers with the structure of molecular beacons (hairpin-like structure)

    Journal: Nucleic Acids Research

    doi:

    Effect of the molecular beacon primer on PCR specificity and sensitivity. ( A ) The target was M.tuberculosis DNA. All reaction mixtures contained 0.5 µg of human placental DNA. Gel electrophoresis in 2% agarose of the amplification products. Lane 1, DNA size marker (100 bp ladder), the prominent band is at 500 bp; lanes 2–4 and 8–10, reverse primer as molecular beacon; lanes 5–7 and 11–13, linear reverse primer; lanes 2, 5, 8 and 11, ∼1000 genome copies; lanes 3, 6, 9 and 12, 100 copies; lanes 4, 7, 10 and 13, 10 copies; lanes 2–7, platinum Taq DNA polymerase; lanes 8–13, normal Taq DNA polymerase. ( B ) Detection of M.tuberculosis DNA in clinical sample by the described technique. Gel electrophoresis in 2% agarose of the amplification products. Lane 1, DNA size marker (100 bp ladder); lanes 2–4 and 8–10, reverse primer as molecular beacon; lanes 5–7 and 11–13, linear reverse primer; lanes 2, 5, 8 and 11, DNA sample from sputum; lanes 3, 6, 9 and 12, ∼1000 genome copies; lanes 4, 7, 10 and 13, ∼100 genome copies; lanes 2–7, platinum Taq DNA polymerase; lanes 8–13, normal Taq DNA polymerase.
    Figure Legend Snippet: Effect of the molecular beacon primer on PCR specificity and sensitivity. ( A ) The target was M.tuberculosis DNA. All reaction mixtures contained 0.5 µg of human placental DNA. Gel electrophoresis in 2% agarose of the amplification products. Lane 1, DNA size marker (100 bp ladder), the prominent band is at 500 bp; lanes 2–4 and 8–10, reverse primer as molecular beacon; lanes 5–7 and 11–13, linear reverse primer; lanes 2, 5, 8 and 11, ∼1000 genome copies; lanes 3, 6, 9 and 12, 100 copies; lanes 4, 7, 10 and 13, 10 copies; lanes 2–7, platinum Taq DNA polymerase; lanes 8–13, normal Taq DNA polymerase. ( B ) Detection of M.tuberculosis DNA in clinical sample by the described technique. Gel electrophoresis in 2% agarose of the amplification products. Lane 1, DNA size marker (100 bp ladder); lanes 2–4 and 8–10, reverse primer as molecular beacon; lanes 5–7 and 11–13, linear reverse primer; lanes 2, 5, 8 and 11, DNA sample from sputum; lanes 3, 6, 9 and 12, ∼1000 genome copies; lanes 4, 7, 10 and 13, ∼100 genome copies; lanes 2–7, platinum Taq DNA polymerase; lanes 8–13, normal Taq DNA polymerase.

    Techniques Used: Polymerase Chain Reaction, DNA Gel Electrophoresis, Amplification, Marker, Nucleic Acid Electrophoresis

    Effect of the molecular beacon primer on the PCR specificity and sensitivity. The target was human DNA (exon 4 of p53 gene). Gel electrophoresis in 2% agarose of the amplification products. Lane 1, DNA size marker (100 bp ladder), the prominent band is 500 bp; lanes 2, 3 and 6, 7, reverse primer as molecular beacon; lanes 4, 5 and 8, 9, linear reverse primer; lanes 2, 4, 6 and 8, ∼1000 genome copies; lanes 3, 5, 7 and 9, ∼100 genome copies; lanes 2–5, platinum Taq DNA polymerase; lanes 8 and 9, usual Taq DNA polymerase. The platinum Taq DNA polymerase gave good amplification from 100 000+ genome copies (not shown).
    Figure Legend Snippet: Effect of the molecular beacon primer on the PCR specificity and sensitivity. The target was human DNA (exon 4 of p53 gene). Gel electrophoresis in 2% agarose of the amplification products. Lane 1, DNA size marker (100 bp ladder), the prominent band is 500 bp; lanes 2, 3 and 6, 7, reverse primer as molecular beacon; lanes 4, 5 and 8, 9, linear reverse primer; lanes 2, 4, 6 and 8, ∼1000 genome copies; lanes 3, 5, 7 and 9, ∼100 genome copies; lanes 2–5, platinum Taq DNA polymerase; lanes 8 and 9, usual Taq DNA polymerase. The platinum Taq DNA polymerase gave good amplification from 100 000+ genome copies (not shown).

    Techniques Used: Polymerase Chain Reaction, Nucleic Acid Electrophoresis, Amplification, Marker

    12) Product Images from "Fidelity of DNA polymerases in the detection of intraindividual variation of mitochondrial DNA"

    Article Title: Fidelity of DNA polymerases in the detection of intraindividual variation of mitochondrial DNA

    Journal: Mitochondrial DNA. Part B, Resources

    doi: 10.1080/23802359.2019.1697188

    The six intraindividual (see aside legend) haplotype networks obtained from COI sequences cloned of Bombus morio (GenBank accession: MK994547-MK994748). COI amplifications were performed using two different polymerases: Platinum Taq DNA Polymerase (A–F) and Q5 DNA polymerase (G–L). Circles represent intraindividual haplotypes, and the size is proportional to their frequency. Crossbars indicate number of nucleotide substitution. Black squares represent a missing intermediate haplotype.
    Figure Legend Snippet: The six intraindividual (see aside legend) haplotype networks obtained from COI sequences cloned of Bombus morio (GenBank accession: MK994547-MK994748). COI amplifications were performed using two different polymerases: Platinum Taq DNA Polymerase (A–F) and Q5 DNA polymerase (G–L). Circles represent intraindividual haplotypes, and the size is proportional to their frequency. Crossbars indicate number of nucleotide substitution. Black squares represent a missing intermediate haplotype.

    Techniques Used: Clone Assay

    13) Product Images from "Improvement of long segment ribosomal PCR amplification for molecular identification of Litylenchus crenatae mccannii associated with beech leaf disease"

    Article Title: Improvement of long segment ribosomal PCR amplification for molecular identification of Litylenchus crenatae mccannii associated with beech leaf disease

    Journal: Journal of Nematology

    doi: 10.21307/jofnem-2020-016

    Long range ribosomal PCR Amplifications of the 3.5 kb target from Fall specimens with Dream Taq ™. M: DNA markers; 1: 104H78; 2: 104H81; 3: 104H82; 4: 104H83; 5: 104H84; 6: 104H85; 7: 104H86; 8: 104H87; 9: 104H88; 10: 104H89; 11: 104H90; NC: negative control. 1-7: Female; 8-11: Male.
    Figure Legend Snippet: Long range ribosomal PCR Amplifications of the 3.5 kb target from Fall specimens with Dream Taq ™. M: DNA markers; 1: 104H78; 2: 104H81; 3: 104H82; 4: 104H83; 5: 104H84; 6: 104H85; 7: 104H86; 8: 104H87; 9: 104H88; 10: 104H89; 11: 104H90; NC: negative control. 1-7: Female; 8-11: Male.

    Techniques Used: Polymerase Chain Reaction, Negative Control

    PCR performance of TaKaRa Ex Taq ® system and PicoMaxx™ High Fidelity PCR System. M: DNA markers; 1 and 5: 104K37; 2 and 6: 104K38; 3 and 7: 104K39; 4 and 8: 104K40. A: 1, 2, 3 and 4: TaKaRa Ex Taq ® system; 5, 6, 7 and 8: PicoMaxx™ High Fidelity PCR System; B: 1, 2, 3 and 4: TaKaRa Ex Taq ® system and Dream Taq ™; 5, 6, 7 and 8: PicoMaxx™ High Fidelity PCR System. NC: negative control, respectively.
    Figure Legend Snippet: PCR performance of TaKaRa Ex Taq ® system and PicoMaxx™ High Fidelity PCR System. M: DNA markers; 1 and 5: 104K37; 2 and 6: 104K38; 3 and 7: 104K39; 4 and 8: 104K40. A: 1, 2, 3 and 4: TaKaRa Ex Taq ® system; 5, 6, 7 and 8: PicoMaxx™ High Fidelity PCR System; B: 1, 2, 3 and 4: TaKaRa Ex Taq ® system and Dream Taq ™; 5, 6, 7 and 8: PicoMaxx™ High Fidelity PCR System. NC: negative control, respectively.

    Techniques Used: Polymerase Chain Reaction, Negative Control

    Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with Dream Taq ™ or/and Pfu in PicoMaxx™ buffer. M: DNA markers; 1, 2, 3 and 4: Dream Taq ™; 5, 6, 7 and 8: Pfu ; 9, 10, 11and 12: Dream Taq ™ and Pfu combined; 1, 5 and 9: 104K29; 2, 6 and 10: 104K30; 3, 7 and 11: 104K31; 4, 8 and 12: negative control (NC), respectively.
    Figure Legend Snippet: Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with Dream Taq ™ or/and Pfu in PicoMaxx™ buffer. M: DNA markers; 1, 2, 3 and 4: Dream Taq ™; 5, 6, 7 and 8: Pfu ; 9, 10, 11and 12: Dream Taq ™ and Pfu combined; 1, 5 and 9: 104K29; 2, 6 and 10: 104K30; 3, 7 and 11: 104K31; 4, 8 and 12: negative control (NC), respectively.

    Techniques Used: Polymerase Chain Reaction, Negative Control

    Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with both Dream Taq ™ and Pfu in manufacturer’s PCR buffers. M: DNA markers; 1: 104K29; 2: 104K30; 3: 104K31; NC: negative control, respectively. A: Dream Taq ™ PCR buffer; B: Pfu PCR buffer.
    Figure Legend Snippet: Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with both Dream Taq ™ and Pfu in manufacturer’s PCR buffers. M: DNA markers; 1: 104K29; 2: 104K30; 3: 104K31; NC: negative control, respectively. A: Dream Taq ™ PCR buffer; B: Pfu PCR buffer.

    Techniques Used: Polymerase Chain Reaction, Negative Control

    Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with TaKaRa Ex Taq ® system. M: DNA markers; 1: 104J54; 2: 104J55; 3: 104J58; 4: 104J59; NC: negative control, respectively. A: Dream Taq ™; B: 18 S locus (1.7 kb) by Dream Taq ™, C: ITS and 28 S loci (1.9 kb) by Dream Taq ™; D: TaKaRa Ex Taq ® system.
    Figure Legend Snippet: Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with TaKaRa Ex Taq ® system. M: DNA markers; 1: 104J54; 2: 104J55; 3: 104J58; 4: 104J59; NC: negative control, respectively. A: Dream Taq ™; B: 18 S locus (1.7 kb) by Dream Taq ™, C: ITS and 28 S loci (1.9 kb) by Dream Taq ™; D: TaKaRa Ex Taq ® system.

    Techniques Used: Polymerase Chain Reaction, Negative Control

    PCR performance of Pfu and Pwo in PicoMaxx™ buffer. M: DNA markers; 1 and 4: 104K37; 2 and 5: 104K38; 3 and 6: 104K39. A: 1, 2 and 3: Dream Taq ™; 4, 5 and 6: Pwo (0.125 μl per reaction). B: 1, 2 and 3: Dream Taq ™ and Pfu ; 4, 5 and 6: Dream Taq ™ and Pwo (0.125 μl per reaction). NC: negative control, respectively. Note: final concentration of Pfu in each reaction was aligned with Pwo and Dream Taq ™ in 0.625 units.
    Figure Legend Snippet: PCR performance of Pfu and Pwo in PicoMaxx™ buffer. M: DNA markers; 1 and 4: 104K37; 2 and 5: 104K38; 3 and 6: 104K39. A: 1, 2 and 3: Dream Taq ™; 4, 5 and 6: Pwo (0.125 μl per reaction). B: 1, 2 and 3: Dream Taq ™ and Pfu ; 4, 5 and 6: Dream Taq ™ and Pwo (0.125 μl per reaction). NC: negative control, respectively. Note: final concentration of Pfu in each reaction was aligned with Pwo and Dream Taq ™ in 0.625 units.

    Techniques Used: Polymerase Chain Reaction, Negative Control, Concentration Assay

    Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with Dream Taq ™ and PicoMaxx™ High Fidelity PCR System. M: DNA markers; 1: 104K25; 2: 104K26; 3: 104K27; 4: 104K28; 5: 104K29; 6: 104K30; 7: 104K31; NC: negative control, respectively. A: 18 S locus (1.7 kb) by Dream Taq ™, B: ITS and 28 S loci (1.9 kb) by Dream Taq ™; C: Dream Taq ™ and PicoMaxx™ High Fidelity PCR System combined.
    Figure Legend Snippet: Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with Dream Taq ™ and PicoMaxx™ High Fidelity PCR System. M: DNA markers; 1: 104K25; 2: 104K26; 3: 104K27; 4: 104K28; 5: 104K29; 6: 104K30; 7: 104K31; NC: negative control, respectively. A: 18 S locus (1.7 kb) by Dream Taq ™, B: ITS and 28 S loci (1.9 kb) by Dream Taq ™; C: Dream Taq ™ and PicoMaxx™ High Fidelity PCR System combined.

    Techniques Used: Polymerase Chain Reaction, Negative Control

    PCR performance of Taq 2000™, Platinum™ Taq and Dream Taq ™. M: DNA markers; 1, 4 and 7: 104N95; 2, 5 and 8: 104N96; 3, 6 and 9: 104N97. 1, 2, 3 and NC by Taq 2000™; 4, 5, 6 and NC by Platinum™ Taq ; 7, 8, 9 and NC by Dream Taq ™, NC: negative control, respectively. A: 3.5 kb target; B: 1.9 kb ITS and 28 S target. Note: final concentration of either Taq 2000™ or Dream Taq ™ in each reaction was aligned with Platinum™ Taq in 1.25 units.
    Figure Legend Snippet: PCR performance of Taq 2000™, Platinum™ Taq and Dream Taq ™. M: DNA markers; 1, 4 and 7: 104N95; 2, 5 and 8: 104N96; 3, 6 and 9: 104N97. 1, 2, 3 and NC by Taq 2000™; 4, 5, 6 and NC by Platinum™ Taq ; 7, 8, 9 and NC by Dream Taq ™, NC: negative control, respectively. A: 3.5 kb target; B: 1.9 kb ITS and 28 S target. Note: final concentration of either Taq 2000™ or Dream Taq ™ in each reaction was aligned with Platinum™ Taq in 1.25 units.

    Techniques Used: Polymerase Chain Reaction, Negative Control, Concentration Assay

    Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with PicoMaxx™ High Fidelity PCR System. M: DNA markers; 1: 104K17; 2: 104K18; 3: 104K19; 4: 104K20; NC: negative control, respectively. A: Dream Taq ™; B: PicoMaxx™ High Fidelity PCR System.
    Figure Legend Snippet: Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with PicoMaxx™ High Fidelity PCR System. M: DNA markers; 1: 104K17; 2: 104K18; 3: 104K19; 4: 104K20; NC: negative control, respectively. A: Dream Taq ™; B: PicoMaxx™ High Fidelity PCR System.

    Techniques Used: Polymerase Chain Reaction, Negative Control

    Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with PicoMaxx™ High Fidelity PCR System. M: DNA markers; 1: 104K25; 2: 104K26; 3: 104K27; 4: 104K28; 5: 104K29; 6: 104K30; 7: 104K31; NC: negative control, respectively. A: Dream Taq ™; B: PicoMaxx™ High Fidelity PCR System.
    Figure Legend Snippet: Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with PicoMaxx™ High Fidelity PCR System. M: DNA markers; 1: 104K25; 2: 104K26; 3: 104K27; 4: 104K28; 5: 104K29; 6: 104K30; 7: 104K31; NC: negative control, respectively. A: Dream Taq ™; B: PicoMaxx™ High Fidelity PCR System.

    Techniques Used: Polymerase Chain Reaction, Negative Control

    14) Product Images from "Molecular Evidence of Orthomyxovirus Presence in Colombian Neotropical Bats"

    Article Title: Molecular Evidence of Orthomyxovirus Presence in Colombian Neotropical Bats

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2022.845546

    Agarose gel electrophoresis of CytB-specific PCR amplified fragments assay. Line 1, 9, and 18: 100 bp DNA ladder (Invitrogen); Line 2: Vampyriscus bidens ; Line 3: Anoura caudifer ; Line 4: Desmodus rotundus ; Line 5: Carollia brevicauda ; Line 6: Carollia castanea ; Line 7: Lonchophylla sp; Line 8: Gardnerycteris crenulatum ; Line 10: Tonatia saurophila ; Line 11: Sturnira tildae ; Line 12: Myotis sp; Line 13: Trachops cirrhosus ; Line 14: Platyrrhinus brachycephalus ; Line 15: Artibeus lituratus ; Line 16: Eptesicus chiriquinus ; Line 17: Mesophylla macconnelli ; Line 19 and 20: Negative controls from MDCK cell line and allantoic fluid from chicken embryonated SPF eggs, respectively.
    Figure Legend Snippet: Agarose gel electrophoresis of CytB-specific PCR amplified fragments assay. Line 1, 9, and 18: 100 bp DNA ladder (Invitrogen); Line 2: Vampyriscus bidens ; Line 3: Anoura caudifer ; Line 4: Desmodus rotundus ; Line 5: Carollia brevicauda ; Line 6: Carollia castanea ; Line 7: Lonchophylla sp; Line 8: Gardnerycteris crenulatum ; Line 10: Tonatia saurophila ; Line 11: Sturnira tildae ; Line 12: Myotis sp; Line 13: Trachops cirrhosus ; Line 14: Platyrrhinus brachycephalus ; Line 15: Artibeus lituratus ; Line 16: Eptesicus chiriquinus ; Line 17: Mesophylla macconnelli ; Line 19 and 20: Negative controls from MDCK cell line and allantoic fluid from chicken embryonated SPF eggs, respectively.

    Techniques Used: Agarose Gel Electrophoresis, Polymerase Chain Reaction, Amplification

    15) Product Images from "Comparison of non-viral methods to genetically modify and enrich populations of primary human corneal endothelial cells"

    Article Title: Comparison of non-viral methods to genetically modify and enrich populations of primary human corneal endothelial cells

    Journal: Molecular Vision

    doi:

    RT-PCR of total RNA of HCECs and human mononuclear cells for COL8A2 and β-actin . Agarose gel shows in lane 1: 1 kb plus ladder, lane 2: COL8A2 with HCECs, lane 3: COL8A2 with HCECs, pretreated with DNase (control to rule out DNA contamination), lane 4: β-actin (housekeeping gene serving as positive control), lane 5: COL8A2 with human mononuclear cells (negative control), lane 6; β-actin with human mononuclear cells (positive control), lane 7: no template (negative control to rule out contamination of reagents), lane 8: PCR with HiFi Taq of HCEC total RNA using COL8A2 primers (negative control to rule out contamination of HCEC total RNA with genomic DNA).
    Figure Legend Snippet: RT-PCR of total RNA of HCECs and human mononuclear cells for COL8A2 and β-actin . Agarose gel shows in lane 1: 1 kb plus ladder, lane 2: COL8A2 with HCECs, lane 3: COL8A2 with HCECs, pretreated with DNase (control to rule out DNA contamination), lane 4: β-actin (housekeeping gene serving as positive control), lane 5: COL8A2 with human mononuclear cells (negative control), lane 6; β-actin with human mononuclear cells (positive control), lane 7: no template (negative control to rule out contamination of reagents), lane 8: PCR with HiFi Taq of HCEC total RNA using COL8A2 primers (negative control to rule out contamination of HCEC total RNA with genomic DNA).

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Agarose Gel Electrophoresis, Positive Control, Negative Control, Polymerase Chain Reaction

    16) Product Images from "Multiplex amplification enabled by selective circularization of large sets of genomic DNA fragments"

    Article Title: Multiplex amplification enabled by selective circularization of large sets of genomic DNA fragments

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gni070

    Selector construct and circularization reaction principles. ( A ) The selector consists of two oligonucleotides, one selector probe of ∼70 nt and one general vector oligonucleotide of 34 nt. The selector probe has two target-specific ends (black) of ∼18 nt complementary to the selected target and a primer-pair motif (gray) of 34 nt. The vector oligonucleotide is complementary to the primer-pair motif (gray). ( B ) The circularization procedure starts with digestion of the DNA to generate targets with defined ends. The digested DNA is then combined with selectors and the mixture is denaturated, allowing the selectors to hybridize to their respective targets. This can be performed in two different ways: (i) a selector probe hybridizes to both ends of the selected target and the ends are connected to the vector by ligation, or (ii) a selector probe hybridizes with one end to the 3′ end of the selected target and the other end to an internal sequence of the target, forming a branched structure that can be cleaved by the endonucleolytic activity of Taq DNA polymerase. Both ends of the selected single-stranded target are now ready for ligation to the vector oligonucleotide, forming a closed circular molecule. The circularization mixture is then exonuclease-treated and finally PCR-amplified using a common primer pair.
    Figure Legend Snippet: Selector construct and circularization reaction principles. ( A ) The selector consists of two oligonucleotides, one selector probe of ∼70 nt and one general vector oligonucleotide of 34 nt. The selector probe has two target-specific ends (black) of ∼18 nt complementary to the selected target and a primer-pair motif (gray) of 34 nt. The vector oligonucleotide is complementary to the primer-pair motif (gray). ( B ) The circularization procedure starts with digestion of the DNA to generate targets with defined ends. The digested DNA is then combined with selectors and the mixture is denaturated, allowing the selectors to hybridize to their respective targets. This can be performed in two different ways: (i) a selector probe hybridizes to both ends of the selected target and the ends are connected to the vector by ligation, or (ii) a selector probe hybridizes with one end to the 3′ end of the selected target and the other end to an internal sequence of the target, forming a branched structure that can be cleaved by the endonucleolytic activity of Taq DNA polymerase. Both ends of the selected single-stranded target are now ready for ligation to the vector oligonucleotide, forming a closed circular molecule. The circularization mixture is then exonuclease-treated and finally PCR-amplified using a common primer pair.

    Techniques Used: Construct, Plasmid Preparation, Ligation, Sequencing, Activity Assay, Polymerase Chain Reaction, Amplification

    17) Product Images from "The chemokine receptor homologue encoded by US27 of human cytomegalovirus is heavily glycosylated and is present in infected human foreskin fibroblasts and enveloped virus particles"

    Article Title: The chemokine receptor homologue encoded by US27 of human cytomegalovirus is heavily glycosylated and is present in infected human foreskin fibroblasts and enveloped virus particles

    Journal: Virus research

    doi: 10.1016/j.virusres.2006.08.003

    (A) N ), to show that RT-PCR and PCR did not fail because of poor enzyme cycling conditions. One-twentieth of the RNA isolated from virus inoculum was used for each experiment. Samples were amplified in the presence of an RT/ Taq DNA polymerase mix (RT-PCR, lanes 11–13 and 17–19) or Platinum Taq ), stained with ethidium bromide, and photographed on a UV transilluminator as part of a Bio-Rad Molecular Imager Gel Doc XR system. Markers (1-kb markers or PCR markers, both from Promega) run in adjacent lanes allowed characterization of the 527-bp US27 PCR product and the 166-bp UL109 PCR product.
    Figure Legend Snippet: (A) N ), to show that RT-PCR and PCR did not fail because of poor enzyme cycling conditions. One-twentieth of the RNA isolated from virus inoculum was used for each experiment. Samples were amplified in the presence of an RT/ Taq DNA polymerase mix (RT-PCR, lanes 11–13 and 17–19) or Platinum Taq ), stained with ethidium bromide, and photographed on a UV transilluminator as part of a Bio-Rad Molecular Imager Gel Doc XR system. Markers (1-kb markers or PCR markers, both from Promega) run in adjacent lanes allowed characterization of the 527-bp US27 PCR product and the 166-bp UL109 PCR product.

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Polymerase Chain Reaction, Isolation, Amplification, Staining

    18) Product Images from "Post-Zygotic and Inter-Individual Structural Genetic Variation in a Presumptive Enhancer Element of the Locus between the IL10Rβ and IFNAR1 Genes"

    Article Title: Post-Zygotic and Inter-Individual Structural Genetic Variation in a Presumptive Enhancer Element of the Locus between the IL10Rβ and IFNAR1 Genes

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0067752

    Variable length of alleles within hypervariable region showing post-zygotic variation. Panel A shows post-zygotic mosaicism in healthy and phenotypically concordant monozygotic twin pair 148341/148342, with five alleles observed in twin 148341, and three alleles present in co-twin 148342. Similarly, panel B displays post-zygotic variation in another monozygotic twin pair 004_01/004_02. In total 5 different alleles are shown on this gel and only one of them is overlapping between both twins. Panel C illustrates post-zygotic mosaicism in breast cancer patient SK58. There are three different alleles in DNA from morphologically normal breast tissue (UM), two alleles in blood cells (BL) and three alleles in primary tumor (PT). In panels A , B and C , Taq DNA polymerase was used for initial PCR amplification from genomic DNA, as indicated by suffix “T” in the ID of each plasmid clone. In panel D , Phusion DNA polymerase confirmed post-zygotic mosaicism in monozygotic twin pair 148341/148342, as indicated by suffix “Ph” in the ID of each plasmid clone. The length of inserts in all plasmid clones was estimated after EcoRI digestion releasing the insert, and using 1% agarose gel. BL, PT and UM indicate peripheral blood DNA, primary breast tumor and healthy morphologically normal breast tissue from a patient affected with breast cancer, respectively.
    Figure Legend Snippet: Variable length of alleles within hypervariable region showing post-zygotic variation. Panel A shows post-zygotic mosaicism in healthy and phenotypically concordant monozygotic twin pair 148341/148342, with five alleles observed in twin 148341, and three alleles present in co-twin 148342. Similarly, panel B displays post-zygotic variation in another monozygotic twin pair 004_01/004_02. In total 5 different alleles are shown on this gel and only one of them is overlapping between both twins. Panel C illustrates post-zygotic mosaicism in breast cancer patient SK58. There are three different alleles in DNA from morphologically normal breast tissue (UM), two alleles in blood cells (BL) and three alleles in primary tumor (PT). In panels A , B and C , Taq DNA polymerase was used for initial PCR amplification from genomic DNA, as indicated by suffix “T” in the ID of each plasmid clone. In panel D , Phusion DNA polymerase confirmed post-zygotic mosaicism in monozygotic twin pair 148341/148342, as indicated by suffix “Ph” in the ID of each plasmid clone. The length of inserts in all plasmid clones was estimated after EcoRI digestion releasing the insert, and using 1% agarose gel. BL, PT and UM indicate peripheral blood DNA, primary breast tumor and healthy morphologically normal breast tissue from a patient affected with breast cancer, respectively.

    Techniques Used: Polymerase Chain Reaction, Amplification, Plasmid Preparation, Clone Assay, Agarose Gel Electrophoresis

    19) Product Images from "Genotyping Echinococcusmultilocularis in Human Alveolar Echinococcosis Patients: An EmsB Microsatellite Analysis"

    Article Title: Genotyping Echinococcusmultilocularis in Human Alveolar Echinococcosis Patients: An EmsB Microsatellite Analysis

    Journal: Pathogens

    doi: 10.3390/pathogens9040282

    Electrophoregrams for the genotyping performed on 14 EchinoRisk samples from three foxes in Multiplex PCR master mix (Qiagen) conditions (MLX A to D) and Platinum Taq DNA polymerase mixture (Invitrogen) conditions (PL) at the Besançon Laboratory (NRC-E) and in AmpliTaq DNA polymerase mixture (Applied Biosystems) conditions at the Berne Laboratory (IPA). The arrows indicate the 2 bp shift observed between the results obtained with the ABI PRISM 3100 Genetic Analyzer and the Applied Biosystems 3130 Genetic Analyzer.
    Figure Legend Snippet: Electrophoregrams for the genotyping performed on 14 EchinoRisk samples from three foxes in Multiplex PCR master mix (Qiagen) conditions (MLX A to D) and Platinum Taq DNA polymerase mixture (Invitrogen) conditions (PL) at the Besançon Laboratory (NRC-E) and in AmpliTaq DNA polymerase mixture (Applied Biosystems) conditions at the Berne Laboratory (IPA). The arrows indicate the 2 bp shift observed between the results obtained with the ABI PRISM 3100 Genetic Analyzer and the Applied Biosystems 3130 Genetic Analyzer.

    Techniques Used: Multiplex Assay, Polymerase Chain Reaction, Indirect Immunoperoxidase Assay

    20) Product Images from "Comparative Methods to Improve the Detection of BRAF V600 Mutations in Highly Pigmented Melanoma Specimens"

    Article Title: Comparative Methods to Improve the Detection of BRAF V600 Mutations in Highly Pigmented Melanoma Specimens

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0158698

    Removal of the inhibitory effects of melanin on PCR amplification. (A) Increasing concentrations of synthetic melanin were added to DNA extracted from 1676 melanoma cell lines. (B) Increasing concentrations of BSA (ng/μl) were added to DNA extracted from cultured cells containing 40 or 80 ng/μl of melanin. (C) The effect of diluting DNA assessed in the presence of 40 ng/μl of melanin and either 1U or 2U of Taq polymerase. (D) NucleoSpin ® gDNA Clean-up XS Kit used on DNA extracted from cultured cells containing 40 or 80 ng/μl of melanin. PCR amplification of the DNA was monitored on 2% gel agarose electrophoresis with ethidium bromide staining. MW, molecular weight markers.
    Figure Legend Snippet: Removal of the inhibitory effects of melanin on PCR amplification. (A) Increasing concentrations of synthetic melanin were added to DNA extracted from 1676 melanoma cell lines. (B) Increasing concentrations of BSA (ng/μl) were added to DNA extracted from cultured cells containing 40 or 80 ng/μl of melanin. (C) The effect of diluting DNA assessed in the presence of 40 ng/μl of melanin and either 1U or 2U of Taq polymerase. (D) NucleoSpin ® gDNA Clean-up XS Kit used on DNA extracted from cultured cells containing 40 or 80 ng/μl of melanin. PCR amplification of the DNA was monitored on 2% gel agarose electrophoresis with ethidium bromide staining. MW, molecular weight markers.

    Techniques Used: Polymerase Chain Reaction, Amplification, Cell Culture, Electrophoresis, Staining, Molecular Weight

    21) Product Images from "A test of somatic mosaicism in the androgen receptor gene of Canada lynx (Lynx canadensis)"

    Article Title: A test of somatic mosaicism in the androgen receptor gene of Canada lynx (Lynx canadensis)

    Journal: BMC Genetics

    doi: 10.1186/s12863-015-0284-y

    Differential peak morphologies of androgen receptor alleles resulting from DNA dilution and reagent use. Lynx positive control DNA sample amplified with Invitrogen Taq DNA Polymerase and diluted to 1:10 ( a ), 1:20 ( b ), and 1:50 ( c ) ratios with deionized water. Lynx positive control DNA sample amplified with Invitrogen Platinum Taq DNA Polymerase (no dilution necessary) ( d )
    Figure Legend Snippet: Differential peak morphologies of androgen receptor alleles resulting from DNA dilution and reagent use. Lynx positive control DNA sample amplified with Invitrogen Taq DNA Polymerase and diluted to 1:10 ( a ), 1:20 ( b ), and 1:50 ( c ) ratios with deionized water. Lynx positive control DNA sample amplified with Invitrogen Platinum Taq DNA Polymerase (no dilution necessary) ( d )

    Techniques Used: Positive Control, Amplification

    22) Product Images from "Beyond the Binding Site: In Vivo Identification of tbx2, smarca5 and wnt5b as Molecular Targets of CNBP during Embryonic Development"

    Article Title: Beyond the Binding Site: In Vivo Identification of tbx2, smarca5 and wnt5b as Molecular Targets of CNBP during Embryonic Development

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0063234

    Consensus sequence for CNBP DNA-binding site. ( A ) A consensus CNBP binding sequence was predicted by applying the MEME program to the 85 and 11 unique sequences obtained from M. musculus and D. rerio libraries, respectively. The logo shown was generated using Weblogo. The 14-nucleotide consensus sequence used for probes design is shown below the logo. Asterisks indicate the six more conserved guanine residues. ( B ) EMSAs were performed using labeled ssDNA probes (C14, C14-mut, C14-comp and dsC14) and increasing concentrations of recombinant CNBP (0.015, 0.050, 0.15, 0.50, 1.50 and 5 µM). Free and shifted probes are indicated by arrows at the left of the gels. K d for C14 is indicated below the gel.
    Figure Legend Snippet: Consensus sequence for CNBP DNA-binding site. ( A ) A consensus CNBP binding sequence was predicted by applying the MEME program to the 85 and 11 unique sequences obtained from M. musculus and D. rerio libraries, respectively. The logo shown was generated using Weblogo. The 14-nucleotide consensus sequence used for probes design is shown below the logo. Asterisks indicate the six more conserved guanine residues. ( B ) EMSAs were performed using labeled ssDNA probes (C14, C14-mut, C14-comp and dsC14) and increasing concentrations of recombinant CNBP (0.015, 0.050, 0.15, 0.50, 1.50 and 5 µM). Free and shifted probes are indicated by arrows at the left of the gels. K d for C14 is indicated below the gel.

    Techniques Used: Sequencing, Binding Assay, Generated, Labeling, Recombinant

    23) Product Images from "Selective control of primer usage in multiplex one-step reverse transcription PCR"

    Article Title: Selective control of primer usage in multiplex one-step reverse transcription PCR

    Journal: BMC Molecular Biology

    doi: 10.1186/1471-2199-10-113

    Real-time one-step RT-PCR evaluation of cDNA priming strategies and the effect of unbalanced target concentrations . A) Real-time one-step RT-PCR evaluation of the effect of different RT primers on quantification of RNA expression in singleplex and in triplex. Reactions employed either an oligo(dT) 18 RT primer, a random decamer RT primer, or a combination of oligo(dT) 18 and random decamer RT primers for cDNA synthesis. In addition, each one-step RT-PCR protocol employed Taq DNA polymerase, M-MLV RT, 0.8 μg of human thymus total RNA, and CleanAmp™ Precision PCR primers. Reactions were performed in triplicate. B) Real-time one-step RT-PCR evaluation of triplex one-step RT-PCR amplifications using different custom prepared mixes containing three RNA standards in different ratios. The relative abundance for each mixture A through H is represented in the following format: (X:Y:Z), where the copies of the ABCA5 RNA standard is present at 10^X copies, the ABCA6 RNA standard is present at 10^Y copies, and the ABCA7 RNA standard is present at 10^Z copies. The observed copy number for each reaction, which was performed in triplicate, was obtained by extrapolation of the Cq to a standard curve for the ABCA5, ABCA6, and ABCA7 RNA standards. The resultant data for each RNA sample was normalized to ABCA7 and was plotted graphically.
    Figure Legend Snippet: Real-time one-step RT-PCR evaluation of cDNA priming strategies and the effect of unbalanced target concentrations . A) Real-time one-step RT-PCR evaluation of the effect of different RT primers on quantification of RNA expression in singleplex and in triplex. Reactions employed either an oligo(dT) 18 RT primer, a random decamer RT primer, or a combination of oligo(dT) 18 and random decamer RT primers for cDNA synthesis. In addition, each one-step RT-PCR protocol employed Taq DNA polymerase, M-MLV RT, 0.8 μg of human thymus total RNA, and CleanAmp™ Precision PCR primers. Reactions were performed in triplicate. B) Real-time one-step RT-PCR evaluation of triplex one-step RT-PCR amplifications using different custom prepared mixes containing three RNA standards in different ratios. The relative abundance for each mixture A through H is represented in the following format: (X:Y:Z), where the copies of the ABCA5 RNA standard is present at 10^X copies, the ABCA6 RNA standard is present at 10^Y copies, and the ABCA7 RNA standard is present at 10^Z copies. The observed copy number for each reaction, which was performed in triplicate, was obtained by extrapolation of the Cq to a standard curve for the ABCA5, ABCA6, and ABCA7 RNA standards. The resultant data for each RNA sample was normalized to ABCA7 and was plotted graphically.

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, RNA Expression, Polymerase Chain Reaction

    One-step RT-PCR evaluation of unmodified and thermolabile CleanAmp™ Precision primers to amplify three different targets of ABCA transporters in singleplex, duplex, and triplex amplifications . For each gene of interest (ABCA5, ABCA6, and ABCA7), the PCR primers were unmodified or contained CleanAmp™ Precision modifications. Reverse transcription utilized an oligo(dT) 18 primer. Reactions contained Taq DNA polymerase, the appropriate reverse transcriptase, and 0.82 μg of human trachea total RNA. A) Reactions employed M-MLV reverse transcriptase and utilized an RT extension temperature of 42°C. B) Reactions employed SuperScript ® III reverse transcriptase (SSIII RT) and utilized an RT extension temperature of 55°C.
    Figure Legend Snippet: One-step RT-PCR evaluation of unmodified and thermolabile CleanAmp™ Precision primers to amplify three different targets of ABCA transporters in singleplex, duplex, and triplex amplifications . For each gene of interest (ABCA5, ABCA6, and ABCA7), the PCR primers were unmodified or contained CleanAmp™ Precision modifications. Reverse transcription utilized an oligo(dT) 18 primer. Reactions contained Taq DNA polymerase, the appropriate reverse transcriptase, and 0.82 μg of human trachea total RNA. A) Reactions employed M-MLV reverse transcriptase and utilized an RT extension temperature of 42°C. B) Reactions employed SuperScript ® III reverse transcriptase (SSIII RT) and utilized an RT extension temperature of 55°C.

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Polymerase Chain Reaction

    Hot Start DNA polymerase evaluation in triplex one-step RT-PCR amplification of ABCA5, ABCA6 and ABCA7 targets . Reactions contained 0.82 μg of human trachea total RNA and an unmodified oligo(dT) 18 RT primer. The PCR primers were either unmodified, or contained CleanAmp™ Precision modifications. These reactions contained one of the following DNA polymerases: Taq , Platinum ® Taq , or AmpliTaq Gold ® and one of the following reverse transcriptases: M-MLV or SSIII. Reactions with M-MLV were incubated at 42°C, while reactions with SSIII were incubated at 55°C.
    Figure Legend Snippet: Hot Start DNA polymerase evaluation in triplex one-step RT-PCR amplification of ABCA5, ABCA6 and ABCA7 targets . Reactions contained 0.82 μg of human trachea total RNA and an unmodified oligo(dT) 18 RT primer. The PCR primers were either unmodified, or contained CleanAmp™ Precision modifications. These reactions contained one of the following DNA polymerases: Taq , Platinum ® Taq , or AmpliTaq Gold ® and one of the following reverse transcriptases: M-MLV or SSIII. Reactions with M-MLV were incubated at 42°C, while reactions with SSIII were incubated at 55°C.

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Amplification, Polymerase Chain Reaction, Incubation

    Singleplex and triplex real-time one-step RT-PCR detection of ABCA5, ABCA6, and ABCA7 in three different tissues . Reactions, which were performed in triplicate, contained M-MLV reverse transcriptase, an unmodified oligo(dT) 18 primer, Taq DNA polymerase and CleanAmp™ Precision PCR primers for the ABCA5, ABCA6 and ABCA7 genes. A standard curve for ABCA5, ABCA6, and ABCA7 was determined by employing ~10 1 to ~10 8 copies of the appropriate RNA standard. Each of the three human total RNA tissue samples (brain (0.78 μg), thymus (0.8 μg), and trachea (0.82 μg)) was amplified in singleplex and triplex format for detection of ABCA5, ABCA6, and ABCA7. The number of copies of each target in a given tissue was determined by extrapolating the resultant Cq values to the standard curve and normalizing the resultant values to the micrograms of input total RNA. A) The relative number of copies per microgram and standard deviation for each target in brain, thymus, and trachea total RNA is represented in a bar graph, which displays the results for singleplex and triplex amplifications. B) The corresponding agarose gel analysis of the three tissue samples amplified in singleplex and in triplex.
    Figure Legend Snippet: Singleplex and triplex real-time one-step RT-PCR detection of ABCA5, ABCA6, and ABCA7 in three different tissues . Reactions, which were performed in triplicate, contained M-MLV reverse transcriptase, an unmodified oligo(dT) 18 primer, Taq DNA polymerase and CleanAmp™ Precision PCR primers for the ABCA5, ABCA6 and ABCA7 genes. A standard curve for ABCA5, ABCA6, and ABCA7 was determined by employing ~10 1 to ~10 8 copies of the appropriate RNA standard. Each of the three human total RNA tissue samples (brain (0.78 μg), thymus (0.8 μg), and trachea (0.82 μg)) was amplified in singleplex and triplex format for detection of ABCA5, ABCA6, and ABCA7. The number of copies of each target in a given tissue was determined by extrapolating the resultant Cq values to the standard curve and normalizing the resultant values to the micrograms of input total RNA. A) The relative number of copies per microgram and standard deviation for each target in brain, thymus, and trachea total RNA is represented in a bar graph, which displays the results for singleplex and triplex amplifications. B) The corresponding agarose gel analysis of the three tissue samples amplified in singleplex and in triplex.

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Polymerase Chain Reaction, Amplification, Standard Deviation, Agarose Gel Electrophoresis

    Evaluation of M-MLV and SSIII reverse transcriptases in multiplex one-step RT-PCR amplification of up to five targets . The amplification of increasing number of targets was evaluated by using either M-MLV RT (42°C) or SSIII RT (55°C). Reactions contained an oligo(dT) 18 primer, 0.82 μg of human trachea total RNA, CleanAmp™ Precision primers, and Taq DNA polymerase.
    Figure Legend Snippet: Evaluation of M-MLV and SSIII reverse transcriptases in multiplex one-step RT-PCR amplification of up to five targets . The amplification of increasing number of targets was evaluated by using either M-MLV RT (42°C) or SSIII RT (55°C). Reactions contained an oligo(dT) 18 primer, 0.82 μg of human trachea total RNA, CleanAmp™ Precision primers, and Taq DNA polymerase.

    Techniques Used: Multiplex Assay, Reverse Transcription Polymerase Chain Reaction, Amplification

    24) Product Images from "Development of a reverse transcription droplet digital PCR (RT-ddPCR) assay for sensitive detection of simian immunodeficiency virus (SIV)"

    Article Title: Development of a reverse transcription droplet digital PCR (RT-ddPCR) assay for sensitive detection of simian immunodeficiency virus (SIV)

    Journal: Virology Journal

    doi: 10.1186/s12985-021-01503-5

    One step RT-ddPCR test. One step RT-ddPCR was performed as described in “Materials and Methods” using the SuperScript III One-Step RT-PCR System with Platinum Taq DNA Polymerase. Assay/sample combinations tested were: SIV single plex assay with buffer ( a ) or SIV RNA standard spike ( b ), SIV single plex assay with 1 μg Rhesus macaque RNA background ( c ) or SIV RNA standard spiked in 1 μg Rhesus macaque RNA background ( d ), SIV and CCR5 duplex assay with buffer ( e ) or SIV RNA standard spike ( f ), and SIV and CCR5 duplex assay with 1 μg Rhesus macaque RNA background ( g ) or with SIV RNA standard spiked in 1 μg Rhesus macaque RNA background ( h ). Detailed experimental conditions are listed in Table 1 . Note that in all panels there were background signals in putative SIV target signal region. Quantitation was not done due to background signals
    Figure Legend Snippet: One step RT-ddPCR test. One step RT-ddPCR was performed as described in “Materials and Methods” using the SuperScript III One-Step RT-PCR System with Platinum Taq DNA Polymerase. Assay/sample combinations tested were: SIV single plex assay with buffer ( a ) or SIV RNA standard spike ( b ), SIV single plex assay with 1 μg Rhesus macaque RNA background ( c ) or SIV RNA standard spiked in 1 μg Rhesus macaque RNA background ( d ), SIV and CCR5 duplex assay with buffer ( e ) or SIV RNA standard spike ( f ), and SIV and CCR5 duplex assay with 1 μg Rhesus macaque RNA background ( g ) or with SIV RNA standard spiked in 1 μg Rhesus macaque RNA background ( h ). Detailed experimental conditions are listed in Table 1 . Note that in all panels there were background signals in putative SIV target signal region. Quantitation was not done due to background signals

    Techniques Used: Reverse Transcription Polymerase Chain Reaction, Plex Assay, Quantitation Assay

    25) Product Images from "Improvement of long segment ribosomal PCR amplification for molecular identification of Litylenchus crenatae mccannii associated with beech leaf disease"

    Article Title: Improvement of long segment ribosomal PCR amplification for molecular identification of Litylenchus crenatae mccannii associated with beech leaf disease

    Journal: Journal of Nematology

    doi: 10.21307/jofnem-2020-016

    Long range ribosomal PCR Amplifications of the 3.5 kb target from Fall specimens with Dream Taq ™. M: DNA markers; 1: 104H78; 2: 104H81; 3: 104H82; 4: 104H83; 5: 104H84; 6: 104H85; 7: 104H86; 8: 104H87; 9: 104H88; 10: 104H89; 11: 104H90; NC: negative control. 1-7: Female; 8-11: Male.
    Figure Legend Snippet: Long range ribosomal PCR Amplifications of the 3.5 kb target from Fall specimens with Dream Taq ™. M: DNA markers; 1: 104H78; 2: 104H81; 3: 104H82; 4: 104H83; 5: 104H84; 6: 104H85; 7: 104H86; 8: 104H87; 9: 104H88; 10: 104H89; 11: 104H90; NC: negative control. 1-7: Female; 8-11: Male.

    Techniques Used: Polymerase Chain Reaction, Negative Control

    PCR performance of TaKaRa Ex Taq ® system and PicoMaxx™ High Fidelity PCR System. M: DNA markers; 1 and 5: 104K37; 2 and 6: 104K38; 3 and 7: 104K39; 4 and 8: 104K40. A: 1, 2, 3 and 4: TaKaRa Ex Taq ® system; 5, 6, 7 and 8: PicoMaxx™ High Fidelity PCR System; B: 1, 2, 3 and 4: TaKaRa Ex Taq ® system and Dream Taq ™; 5, 6, 7 and 8: PicoMaxx™ High Fidelity PCR System. NC: negative control, respectively.
    Figure Legend Snippet: PCR performance of TaKaRa Ex Taq ® system and PicoMaxx™ High Fidelity PCR System. M: DNA markers; 1 and 5: 104K37; 2 and 6: 104K38; 3 and 7: 104K39; 4 and 8: 104K40. A: 1, 2, 3 and 4: TaKaRa Ex Taq ® system; 5, 6, 7 and 8: PicoMaxx™ High Fidelity PCR System; B: 1, 2, 3 and 4: TaKaRa Ex Taq ® system and Dream Taq ™; 5, 6, 7 and 8: PicoMaxx™ High Fidelity PCR System. NC: negative control, respectively.

    Techniques Used: Polymerase Chain Reaction, Negative Control

    Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with Dream Taq ™ or/and Pfu in PicoMaxx™ buffer. M: DNA markers; 1, 2, 3 and 4: Dream Taq ™; 5, 6, 7 and 8: Pfu ; 9, 10, 11and 12: Dream Taq ™ and Pfu combined; 1, 5 and 9: 104K29; 2, 6 and 10: 104K30; 3, 7 and 11: 104K31; 4, 8 and 12: negative control (NC), respectively.
    Figure Legend Snippet: Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with Dream Taq ™ or/and Pfu in PicoMaxx™ buffer. M: DNA markers; 1, 2, 3 and 4: Dream Taq ™; 5, 6, 7 and 8: Pfu ; 9, 10, 11and 12: Dream Taq ™ and Pfu combined; 1, 5 and 9: 104K29; 2, 6 and 10: 104K30; 3, 7 and 11: 104K31; 4, 8 and 12: negative control (NC), respectively.

    Techniques Used: Polymerase Chain Reaction, Negative Control

    Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with both Dream Taq ™ and Pfu in manufacturer’s PCR buffers. M: DNA markers; 1: 104K29; 2: 104K30; 3: 104K31; NC: negative control, respectively. A: Dream Taq ™ PCR buffer; B: Pfu PCR buffer.
    Figure Legend Snippet: Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with both Dream Taq ™ and Pfu in manufacturer’s PCR buffers. M: DNA markers; 1: 104K29; 2: 104K30; 3: 104K31; NC: negative control, respectively. A: Dream Taq ™ PCR buffer; B: Pfu PCR buffer.

    Techniques Used: Polymerase Chain Reaction, Negative Control

    Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with TaKaRa Ex Taq ® system. M: DNA markers; 1: 104J54; 2: 104J55; 3: 104J58; 4: 104J59; NC: negative control, respectively. A: Dream Taq ™; B: 18 S locus (1.7 kb) by Dream Taq ™, C: ITS and 28 S loci (1.9 kb) by Dream Taq ™; D: TaKaRa Ex Taq ® system.
    Figure Legend Snippet: Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with TaKaRa Ex Taq ® system. M: DNA markers; 1: 104J54; 2: 104J55; 3: 104J58; 4: 104J59; NC: negative control, respectively. A: Dream Taq ™; B: 18 S locus (1.7 kb) by Dream Taq ™, C: ITS and 28 S loci (1.9 kb) by Dream Taq ™; D: TaKaRa Ex Taq ® system.

    Techniques Used: Polymerase Chain Reaction, Negative Control

    PCR performance of Pfu and Pwo in PicoMaxx™ buffer. M: DNA markers; 1 and 4: 104K37; 2 and 5: 104K38; 3 and 6: 104K39. A: 1, 2 and 3: Dream Taq ™; 4, 5 and 6: Pwo (0.125 μl per reaction). B: 1, 2 and 3: Dream Taq ™ and Pfu ; 4, 5 and 6: Dream Taq ™ and Pwo (0.125 μl per reaction). NC: negative control, respectively. Note: final concentration of Pfu in each reaction was aligned with Pwo and Dream Taq ™ in 0.625 units.
    Figure Legend Snippet: PCR performance of Pfu and Pwo in PicoMaxx™ buffer. M: DNA markers; 1 and 4: 104K37; 2 and 5: 104K38; 3 and 6: 104K39. A: 1, 2 and 3: Dream Taq ™; 4, 5 and 6: Pwo (0.125 μl per reaction). B: 1, 2 and 3: Dream Taq ™ and Pfu ; 4, 5 and 6: Dream Taq ™ and Pwo (0.125 μl per reaction). NC: negative control, respectively. Note: final concentration of Pfu in each reaction was aligned with Pwo and Dream Taq ™ in 0.625 units.

    Techniques Used: Polymerase Chain Reaction, Negative Control, Concentration Assay

    Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with Dream Taq ™ and PicoMaxx™ High Fidelity PCR System. M: DNA markers; 1: 104K25; 2: 104K26; 3: 104K27; 4: 104K28; 5: 104K29; 6: 104K30; 7: 104K31; NC: negative control, respectively. A: 18 S locus (1.7 kb) by Dream Taq ™, B: ITS and 28 S loci (1.9 kb) by Dream Taq ™; C: Dream Taq ™ and PicoMaxx™ High Fidelity PCR System combined.
    Figure Legend Snippet: Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with Dream Taq ™ and PicoMaxx™ High Fidelity PCR System. M: DNA markers; 1: 104K25; 2: 104K26; 3: 104K27; 4: 104K28; 5: 104K29; 6: 104K30; 7: 104K31; NC: negative control, respectively. A: 18 S locus (1.7 kb) by Dream Taq ™, B: ITS and 28 S loci (1.9 kb) by Dream Taq ™; C: Dream Taq ™ and PicoMaxx™ High Fidelity PCR System combined.

    Techniques Used: Polymerase Chain Reaction, Negative Control

    PCR performance of Taq 2000™, Platinum™ Taq and Dream Taq ™. M: DNA markers; 1, 4 and 7: 104N95; 2, 5 and 8: 104N96; 3, 6 and 9: 104N97. 1, 2, 3 and NC by Taq 2000™; 4, 5, 6 and NC by Platinum™ Taq ; 7, 8, 9 and NC by Dream Taq ™, NC: negative control, respectively. A: 3.5 kb target; B: 1.9 kb ITS and 28 S target. Note: final concentration of either Taq 2000™ or Dream Taq ™ in each reaction was aligned with Platinum™ Taq in 1.25 units.
    Figure Legend Snippet: PCR performance of Taq 2000™, Platinum™ Taq and Dream Taq ™. M: DNA markers; 1, 4 and 7: 104N95; 2, 5 and 8: 104N96; 3, 6 and 9: 104N97. 1, 2, 3 and NC by Taq 2000™; 4, 5, 6 and NC by Platinum™ Taq ; 7, 8, 9 and NC by Dream Taq ™, NC: negative control, respectively. A: 3.5 kb target; B: 1.9 kb ITS and 28 S target. Note: final concentration of either Taq 2000™ or Dream Taq ™ in each reaction was aligned with Platinum™ Taq in 1.25 units.

    Techniques Used: Polymerase Chain Reaction, Negative Control, Concentration Assay

    Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with PicoMaxx™ High Fidelity PCR System. M: DNA markers; 1: 104K17; 2: 104K18; 3: 104K19; 4: 104K20; NC: negative control, respectively. A: Dream Taq ™; B: PicoMaxx™ High Fidelity PCR System.
    Figure Legend Snippet: Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with PicoMaxx™ High Fidelity PCR System. M: DNA markers; 1: 104K17; 2: 104K18; 3: 104K19; 4: 104K20; NC: negative control, respectively. A: Dream Taq ™; B: PicoMaxx™ High Fidelity PCR System.

    Techniques Used: Polymerase Chain Reaction, Negative Control

    Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with PicoMaxx™ High Fidelity PCR System. M: DNA markers; 1: 104K25; 2: 104K26; 3: 104K27; 4: 104K28; 5: 104K29; 6: 104K30; 7: 104K31; NC: negative control, respectively. A: Dream Taq ™; B: PicoMaxx™ High Fidelity PCR System.
    Figure Legend Snippet: Long range ribosomal PCR Amplifications of the 3.5 kb target from Summer specimens with PicoMaxx™ High Fidelity PCR System. M: DNA markers; 1: 104K25; 2: 104K26; 3: 104K27; 4: 104K28; 5: 104K29; 6: 104K30; 7: 104K31; NC: negative control, respectively. A: Dream Taq ™; B: PicoMaxx™ High Fidelity PCR System.

    Techniques Used: Polymerase Chain Reaction, Negative Control

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  • 99
    Thermo Fisher high fidelity platinum pfx dna polymerase
    Polyglutamine-mediated interaction influences CIZ1 assembly structure at Xi A) Example images showing endogenous CIZ1 (C-term, green) and H2AK119ub (red) in female WT PEFs at p3, without (left) and with (right) incubation with polyglutamine aggregation inhibitor C2-8 (inset) for 24 hours. <t>DNA</t> is blue. Bar is 5 microns. Below, example nucleus with i) lamina-associated ribbon-like CIZ1 assembly. B) Box and whisker plot showing area of CIZ1 assemblies (green signal), calculated as a proportion of nuclear area (blue signal), generated using image masks in FIJI (below). Data is representative of two experiments with independent WT primary cell isolates. C) Histogram showing that the frequency of CIZ1 assemblies remains unaffected by C2-8. D) Left, schematic of transgenes used to create doxycycline (dox)-inducible expression of full-length GFP-CIZ1 in CIZ1 null mice derived cells ( Ridings-Figueroa et al., 2017 ). Tet-responsive element (TRE), CMV promoter (CMV), and reverse tetracycline transcriptional activator (rtTA). Histogram shows frequency of globular or ribbon-like GFP-CIZ1 assemblies (green), or absence of assemblies (grey) when cycling CIZ1 null PEFs at p3 were exposed to the indicated concentrations of C2-8 throughout a 48 hour induction period. Right, example images showing accumulation of GFP-CIZ1 (green) at Xi, either as a typical globular structure or an elongated ribbon-like structure associated with the nuclear lamina. Nuclei are counterstained for lamin B (red), DNA is blue, bar is 5 microns. Results are representative of three experiments. E) Histogram shows the proportion of GFP-CIZ1 assemblies that co-stain for H2AK119ub or H3K27me3 in female CIZ1 null PEFs (p3), in the presence or absence of C2-8, where ribbons are shown in pink and globular assemblies in red. Grey, no mark. Error bars are SEM from three replicates, ns, not significant, indicating that modification of Xi chromatin persists in assemblies whose structure is affected by C2-8. Comparisons are by t-test. Right, example images showing induced CIZ1-assemblies (green) and the indicated chromatin marks in red, DNA is blue. Bar is 5 microns. Far right, individual channels as indicated. F) Example images showing retention of <t>Xist</t> (red) upon induction of GFP-CIZ1 and the formation of de novo CIZ1 assemblies (green), in the absence and (two examples) presence of C2-8. Far right, high magnification, separate views of CIZ1 and Xist in grey scale. Far left, example WT cell showing normal Xist cloud.
    High Fidelity Platinum Pfx Dna Polymerase, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 437 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher platinum taq dna polymerase
    Workflow of reverse transcription and amplification of cfmRNA. cfmRNA, spiked-in with luciferase RNA control, was reverse transcript with Superscript™ III Reverse Transcriptase (Invitrogen, Cat no. 18080044). The product was amplified with rhPCR primers and Platinum™ <t>Taq</t> <t>DNA</t> Polymerase (Invitrogen, Cat no. 10966) using emulsion and CoT PCR. The residual primers were removed with Exonuclease I (New England Biolabs, Cat no. M0293). Amplified products were used for qPCR quantification and sequencing.
    Platinum Taq Dna Polymerase, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Polyglutamine-mediated interaction influences CIZ1 assembly structure at Xi A) Example images showing endogenous CIZ1 (C-term, green) and H2AK119ub (red) in female WT PEFs at p3, without (left) and with (right) incubation with polyglutamine aggregation inhibitor C2-8 (inset) for 24 hours. DNA is blue. Bar is 5 microns. Below, example nucleus with i) lamina-associated ribbon-like CIZ1 assembly. B) Box and whisker plot showing area of CIZ1 assemblies (green signal), calculated as a proportion of nuclear area (blue signal), generated using image masks in FIJI (below). Data is representative of two experiments with independent WT primary cell isolates. C) Histogram showing that the frequency of CIZ1 assemblies remains unaffected by C2-8. D) Left, schematic of transgenes used to create doxycycline (dox)-inducible expression of full-length GFP-CIZ1 in CIZ1 null mice derived cells ( Ridings-Figueroa et al., 2017 ). Tet-responsive element (TRE), CMV promoter (CMV), and reverse tetracycline transcriptional activator (rtTA). Histogram shows frequency of globular or ribbon-like GFP-CIZ1 assemblies (green), or absence of assemblies (grey) when cycling CIZ1 null PEFs at p3 were exposed to the indicated concentrations of C2-8 throughout a 48 hour induction period. Right, example images showing accumulation of GFP-CIZ1 (green) at Xi, either as a typical globular structure or an elongated ribbon-like structure associated with the nuclear lamina. Nuclei are counterstained for lamin B (red), DNA is blue, bar is 5 microns. Results are representative of three experiments. E) Histogram shows the proportion of GFP-CIZ1 assemblies that co-stain for H2AK119ub or H3K27me3 in female CIZ1 null PEFs (p3), in the presence or absence of C2-8, where ribbons are shown in pink and globular assemblies in red. Grey, no mark. Error bars are SEM from three replicates, ns, not significant, indicating that modification of Xi chromatin persists in assemblies whose structure is affected by C2-8. Comparisons are by t-test. Right, example images showing induced CIZ1-assemblies (green) and the indicated chromatin marks in red, DNA is blue. Bar is 5 microns. Far right, individual channels as indicated. F) Example images showing retention of Xist (red) upon induction of GFP-CIZ1 and the formation of de novo CIZ1 assemblies (green), in the absence and (two examples) presence of C2-8. Far right, high magnification, separate views of CIZ1 and Xist in grey scale. Far left, example WT cell showing normal Xist cloud.

    Journal: bioRxiv

    Article Title: A polyglutamine domain is required for de novo CIZ1 assembly formation at the inactive X chromosome

    doi: 10.1101/2020.11.10.376558

    Figure Lengend Snippet: Polyglutamine-mediated interaction influences CIZ1 assembly structure at Xi A) Example images showing endogenous CIZ1 (C-term, green) and H2AK119ub (red) in female WT PEFs at p3, without (left) and with (right) incubation with polyglutamine aggregation inhibitor C2-8 (inset) for 24 hours. DNA is blue. Bar is 5 microns. Below, example nucleus with i) lamina-associated ribbon-like CIZ1 assembly. B) Box and whisker plot showing area of CIZ1 assemblies (green signal), calculated as a proportion of nuclear area (blue signal), generated using image masks in FIJI (below). Data is representative of two experiments with independent WT primary cell isolates. C) Histogram showing that the frequency of CIZ1 assemblies remains unaffected by C2-8. D) Left, schematic of transgenes used to create doxycycline (dox)-inducible expression of full-length GFP-CIZ1 in CIZ1 null mice derived cells ( Ridings-Figueroa et al., 2017 ). Tet-responsive element (TRE), CMV promoter (CMV), and reverse tetracycline transcriptional activator (rtTA). Histogram shows frequency of globular or ribbon-like GFP-CIZ1 assemblies (green), or absence of assemblies (grey) when cycling CIZ1 null PEFs at p3 were exposed to the indicated concentrations of C2-8 throughout a 48 hour induction period. Right, example images showing accumulation of GFP-CIZ1 (green) at Xi, either as a typical globular structure or an elongated ribbon-like structure associated with the nuclear lamina. Nuclei are counterstained for lamin B (red), DNA is blue, bar is 5 microns. Results are representative of three experiments. E) Histogram shows the proportion of GFP-CIZ1 assemblies that co-stain for H2AK119ub or H3K27me3 in female CIZ1 null PEFs (p3), in the presence or absence of C2-8, where ribbons are shown in pink and globular assemblies in red. Grey, no mark. Error bars are SEM from three replicates, ns, not significant, indicating that modification of Xi chromatin persists in assemblies whose structure is affected by C2-8. Comparisons are by t-test. Right, example images showing induced CIZ1-assemblies (green) and the indicated chromatin marks in red, DNA is blue. Bar is 5 microns. Far right, individual channels as indicated. F) Example images showing retention of Xist (red) upon induction of GFP-CIZ1 and the formation of de novo CIZ1 assemblies (green), in the absence and (two examples) presence of C2-8. Far right, high magnification, separate views of CIZ1 and Xist in grey scale. Far left, example WT cell showing normal Xist cloud.

    Article Snippet: In vitro transcription of DIG labelled probes DNA templates used for in vitro transcription of mouse Xist RNA were amplified from sequence verified plasmid pCMV-Xist-PA ( ) (Addgene 26760), containing the murine Xist gene using high-fidelity Platinum® pfx DNA polymerase (Invitrogen™).

    Techniques: Incubation, Whisker Assay, Generated, Expressing, Mouse Assay, Derivative Assay, Staining, Modification

    Three alleles of AtECA3 T-DNA insertional mutants were identified. A, Genomic structure of AtECA3 showing T-DNA insertion sites of eca3-1b , eca3-4 , and eca3-5 . The insertions were verified by left border sequencing. Bases in uppercase and lowercase letters

    Journal:

    Article Title: A Distinct Endosomal Ca2+/Mn2+ Pump Affects Root Growth through the Secretory Process 1 Pump Affects Root Growth through the Secretory Process 1 [C] Pump Affects Root Growth through the Secretory Process 1 [C] [W] Pump Affects Root Growth through the Secretory Process 1 [C] [W] [OA]

    doi: 10.1104/pp.108.119909

    Figure Lengend Snippet: Three alleles of AtECA3 T-DNA insertional mutants were identified. A, Genomic structure of AtECA3 showing T-DNA insertion sites of eca3-1b , eca3-4 , and eca3-5 . The insertions were verified by left border sequencing. Bases in uppercase and lowercase letters

    Article Snippet: For Gateway cloning, the AtECA3 cDNA was amplified by PCR using Platinum Pfx DNA polymerase (Invitrogen) and primers ECA3-Gf and ECA3-Gr.

    Techniques: Sequencing

    DNA sequences tested in EMSA with P22 Xis. All experiments were performed with annealed, double-stranded DNA oligonucleotides. Quantitation was performed as described in Materials and Methods. The DNA sequence protected by the P22 Xis DNase I footprint is at the top. For orientation, the direct repeats found in the footprint sequences have been boxed (from left to right, X1 to X4).

    Journal: Journal of Bacteriology

    Article Title: Purification and Characterization of Bacteriophage P22 Xis Protein ▿

    doi: 10.1128/JB.00170-08

    Figure Lengend Snippet: DNA sequences tested in EMSA with P22 Xis. All experiments were performed with annealed, double-stranded DNA oligonucleotides. Quantitation was performed as described in Materials and Methods. The DNA sequence protected by the P22 Xis DNase I footprint is at the top. For orientation, the direct repeats found in the footprint sequences have been boxed (from left to right, X1 to X4).

    Article Snippet: PCR product XISDBS29 was amplified from P22 attP (plasmid pRA114) by using Platinum Pfx DNA polymerase according to the Invitrogen protocol with 32 P-labeled primers 802up and 806do.

    Techniques: Quantitation Assay, Sequencing

    Alignment of attP sites from lambda, P22, L5, HP1, P2, and Tn 916 . The sites are aligned at the right, along the middle of the two Int cleavage sites. DNA core sites are in green and their footprints in yellow. All other Int binding sites not at the point of cleavage are in blue. Xis proposed or known binding recognition sites are shown in red. The Xis footprint-protected bases are marked by the black line below the red recognition site for the strand shown. The second black line below the first represents protected bases for the cDNA strand, which is not shown (HP1 and P2 only). IHF or host factor binding sites are shown in gray. The mIHF protection site in L5 is shown in purple because no recognition sequence near the protection site has been proposed.

    Journal: Journal of Bacteriology

    Article Title: Purification and Characterization of Bacteriophage P22 Xis Protein ▿

    doi: 10.1128/JB.00170-08

    Figure Lengend Snippet: Alignment of attP sites from lambda, P22, L5, HP1, P2, and Tn 916 . The sites are aligned at the right, along the middle of the two Int cleavage sites. DNA core sites are in green and their footprints in yellow. All other Int binding sites not at the point of cleavage are in blue. Xis proposed or known binding recognition sites are shown in red. The Xis footprint-protected bases are marked by the black line below the red recognition site for the strand shown. The second black line below the first represents protected bases for the cDNA strand, which is not shown (HP1 and P2 only). IHF or host factor binding sites are shown in gray. The mIHF protection site in L5 is shown in purple because no recognition sequence near the protection site has been proposed.

    Article Snippet: PCR product XISDBS29 was amplified from P22 attP (plasmid pRA114) by using Platinum Pfx DNA polymerase according to the Invitrogen protocol with 32 P-labeled primers 802up and 806do.

    Techniques: Binding Assay, Immunohistofluorescence, Sequencing

    (A) DNase I footprint of P22 Xis on P22 attP DNA (XISDBS29), top strand. Concentrations of P22 Xis are indicated above the gel. Lanes 0 contain no Xis protein. P22 attP DNA binding sites P1, P2, and P3 are labeled on the left. A protected region, illustrated by the bracket, contains direct repeats X1, X2, X3, and X4. Enhanced cleavages are indicated by the arrows. T+C and G+A ladders are shown in the leftmost two lanes. (B) DNase I footprint of P22 Xis on P22 attP DNA (XISDBS29), bottom strand. Concentrations of P22 Xis are indicated above. Lanes 0 contain no Xis protein. P22 attP DNA binding sites P1, P2, and P3 are labeled on the left. Direct repeats (X1 to X4) are labeled and are located in the area that was clearly protected on the complementary strand. Enhanced cleavages are indicated by the arrows. T+C and G+A ladders are shown for the same DNA fragment (left lanes). Lanes 0 contain no Xis protein. (C) DNase I footprint of P22 Xis on P22 attP (P22attP367, top strand) DNA showing secondary footprint sites. Lanes contain increasing concentrations of P22 Xis as indicated. Lane 0 contains no Xis protein. A DNA sequencing ladder is shown on the right to identify the protected bases, which are represented as A, C, G, and T. To the right is a map of P22 attP . The secondary protected regions cover the C, C′, H, and H′ DNA binding sites. (D) Summary of DNA footprinting results. Bases specifically protected by secondary footprints. The P22 attP DNA binding sites are shown and labeled as follows: light blue, arm-type binding sites P1, P2, and P3; green, core Int binding sites C and C′; and gray, IHF binding sites H and H′. The P22 Xis DNA binding sites protected at low protein concentration are shown by solid bars above the strand. Sites protected by highest concentration of P22 Xis (including secondary sites) are shown by open bars above the strand. Direct repeats in the P22 Xis primary footprint site are shown in red and labeled as X1, X2, X3, and X4. Hypersensitive sites are marked with asterisks.

    Journal: Journal of Bacteriology

    Article Title: Purification and Characterization of Bacteriophage P22 Xis Protein ▿

    doi: 10.1128/JB.00170-08

    Figure Lengend Snippet: (A) DNase I footprint of P22 Xis on P22 attP DNA (XISDBS29), top strand. Concentrations of P22 Xis are indicated above the gel. Lanes 0 contain no Xis protein. P22 attP DNA binding sites P1, P2, and P3 are labeled on the left. A protected region, illustrated by the bracket, contains direct repeats X1, X2, X3, and X4. Enhanced cleavages are indicated by the arrows. T+C and G+A ladders are shown in the leftmost two lanes. (B) DNase I footprint of P22 Xis on P22 attP DNA (XISDBS29), bottom strand. Concentrations of P22 Xis are indicated above. Lanes 0 contain no Xis protein. P22 attP DNA binding sites P1, P2, and P3 are labeled on the left. Direct repeats (X1 to X4) are labeled and are located in the area that was clearly protected on the complementary strand. Enhanced cleavages are indicated by the arrows. T+C and G+A ladders are shown for the same DNA fragment (left lanes). Lanes 0 contain no Xis protein. (C) DNase I footprint of P22 Xis on P22 attP (P22attP367, top strand) DNA showing secondary footprint sites. Lanes contain increasing concentrations of P22 Xis as indicated. Lane 0 contains no Xis protein. A DNA sequencing ladder is shown on the right to identify the protected bases, which are represented as A, C, G, and T. To the right is a map of P22 attP . The secondary protected regions cover the C, C′, H, and H′ DNA binding sites. (D) Summary of DNA footprinting results. Bases specifically protected by secondary footprints. The P22 attP DNA binding sites are shown and labeled as follows: light blue, arm-type binding sites P1, P2, and P3; green, core Int binding sites C and C′; and gray, IHF binding sites H and H′. The P22 Xis DNA binding sites protected at low protein concentration are shown by solid bars above the strand. Sites protected by highest concentration of P22 Xis (including secondary sites) are shown by open bars above the strand. Direct repeats in the P22 Xis primary footprint site are shown in red and labeled as X1, X2, X3, and X4. Hypersensitive sites are marked with asterisks.

    Article Snippet: PCR product XISDBS29 was amplified from P22 attP (plasmid pRA114) by using Platinum Pfx DNA polymerase according to the Invitrogen protocol with 32 P-labeled primers 802up and 806do.

    Techniques: Binding Assay, Labeling, DNA Sequencing, DNA Footprinting, Immunohistofluorescence, Protein Concentration, Concentration Assay

    Workflow of reverse transcription and amplification of cfmRNA. cfmRNA, spiked-in with luciferase RNA control, was reverse transcript with Superscript™ III Reverse Transcriptase (Invitrogen, Cat no. 18080044). The product was amplified with rhPCR primers and Platinum™ Taq DNA Polymerase (Invitrogen, Cat no. 10966) using emulsion and CoT PCR. The residual primers were removed with Exonuclease I (New England Biolabs, Cat no. M0293). Amplified products were used for qPCR quantification and sequencing.

    Journal: Frontiers in Genetics

    Article Title: Non-Invasive Characterization of the Pancreas During Bariatric Surgery via Circulating Pancreatic Specific Cell-free Messenger RNA

    doi: 10.3389/fgene.2021.742496

    Figure Lengend Snippet: Workflow of reverse transcription and amplification of cfmRNA. cfmRNA, spiked-in with luciferase RNA control, was reverse transcript with Superscript™ III Reverse Transcriptase (Invitrogen, Cat no. 18080044). The product was amplified with rhPCR primers and Platinum™ Taq DNA Polymerase (Invitrogen, Cat no. 10966) using emulsion and CoT PCR. The residual primers were removed with Exonuclease I (New England Biolabs, Cat no. M0293). Amplified products were used for qPCR quantification and sequencing.

    Article Snippet: Reverse transcription of cfRNA was performed using Superscript™ III Reverse Transcriptase (Invitrogen, Cat no. 18080044) at 25°C for 5 min, 50°C for 50 min, and enzyme inactivation at 95°C for 3 min. cDNA from reverse transcription was added to the PCR mixture of Platinum™ Taq DNA Polymerase (Invitrogen, Cat no. 10966) with a final concentration of 0.5 μM of rh PCR primers mix and 26 mU of RNase H2 enzyme.

    Techniques: Amplification, Luciferase, RNase H-dependent PCR, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Sequencing