control input dna Search Results


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  • 93
    Zymo Research input control dna
    Input Control Dna, supplied by Zymo Research, used in various techniques. Bioz Stars score: 93/100, based on 72 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    New England Biolabs t4 dna polymerase
    T4 Dna Polymerase, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 99/100, based on 9070 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore control input dna
    Comparison of methylation data between <t>MeDIP/NimbleGen</t> Promoter + CpGi Array and Sequenom MassArray: Stk31 CpGi promoter region . The Stk31 gene region with a CpGi promoter region that is methylated in the liver, but not in ES or testis cells. The box indicates the position of the methylation peak in the liver that overlaps with the CpGi promoter region analyzed by Sequenom. In the epigram of Sequenom methylation analysis shown at the bottom panel, the blue circles indicate 100% methylation, green circles indicate around 50%methylation and yellow circles indicate 0% methylation. The results from biological duplicate samples are shown. The bottom lane (Sequenom Methylation Analysis) shows the results of analysis of testis <t>DNA</t> after in vitro methylation using M. Sss1 methylase (New England Biolabs).
    Control Input Dna, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 146 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher control input dna
    Comparison of methylation data between <t>MeDIP/NimbleGen</t> Promoter + CpGi Array and Sequenom MassArray: Stk31 CpGi promoter region . The Stk31 gene region with a CpGi promoter region that is methylated in the liver, but not in ES or testis cells. The box indicates the position of the methylation peak in the liver that overlaps with the CpGi promoter region analyzed by Sequenom. In the epigram of Sequenom methylation analysis shown at the bottom panel, the blue circles indicate 100% methylation, green circles indicate around 50%methylation and yellow circles indicate 0% methylation. The results from biological duplicate samples are shown. The bottom lane (Sequenom Methylation Analysis) shows the results of analysis of testis <t>DNA</t> after in vitro methylation using M. Sss1 methylase (New England Biolabs).
    Control Input Dna, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 18 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Illumina Inc input dna control
    Comparison of methylation data between <t>MeDIP/NimbleGen</t> Promoter + CpGi Array and Sequenom MassArray: Stk31 CpGi promoter region . The Stk31 gene region with a CpGi promoter region that is methylated in the liver, but not in ES or testis cells. The box indicates the position of the methylation peak in the liver that overlaps with the CpGi promoter region analyzed by Sequenom. In the epigram of Sequenom methylation analysis shown at the bottom panel, the blue circles indicate 100% methylation, green circles indicate around 50%methylation and yellow circles indicate 0% methylation. The results from biological duplicate samples are shown. The bottom lane (Sequenom Methylation Analysis) shows the results of analysis of testis <t>DNA</t> after in vitro methylation using M. Sss1 methylase (New England Biolabs).
    Input Dna Control, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 93/100, based on 19 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Illumina Inc control dna input
    Comparison of methylation data between <t>MeDIP/NimbleGen</t> Promoter + CpGi Array and Sequenom MassArray: Stk31 CpGi promoter region . The Stk31 gene region with a CpGi promoter region that is methylated in the liver, but not in ES or testis cells. The box indicates the position of the methylation peak in the liver that overlaps with the CpGi promoter region analyzed by Sequenom. In the epigram of Sequenom methylation analysis shown at the bottom panel, the blue circles indicate 100% methylation, green circles indicate around 50%methylation and yellow circles indicate 0% methylation. The results from biological duplicate samples are shown. The bottom lane (Sequenom Methylation Analysis) shows the results of analysis of testis <t>DNA</t> after in vitro methylation using M. Sss1 methylase (New England Biolabs).
    Control Dna Input, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 89/100, based on 37 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher input cdna control
    Comparison of methylation data between <t>MeDIP/NimbleGen</t> Promoter + CpGi Array and Sequenom MassArray: Stk31 CpGi promoter region . The Stk31 gene region with a CpGi promoter region that is methylated in the liver, but not in ES or testis cells. The box indicates the position of the methylation peak in the liver that overlaps with the CpGi promoter region analyzed by Sequenom. In the epigram of Sequenom methylation analysis shown at the bottom panel, the blue circles indicate 100% methylation, green circles indicate around 50%methylation and yellow circles indicate 0% methylation. The results from biological duplicate samples are shown. The bottom lane (Sequenom Methylation Analysis) shows the results of analysis of testis <t>DNA</t> after in vitro methylation using M. Sss1 methylase (New England Biolabs).
    Input Cdna Control, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 85/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Millipore control dna
    Comparison of methylation data between <t>MeDIP/NimbleGen</t> Promoter + CpGi Array and Sequenom MassArray: Stk31 CpGi promoter region . The Stk31 gene region with a CpGi promoter region that is methylated in the liver, but not in ES or testis cells. The box indicates the position of the methylation peak in the liver that overlaps with the CpGi promoter region analyzed by Sequenom. In the epigram of Sequenom methylation analysis shown at the bottom panel, the blue circles indicate 100% methylation, green circles indicate around 50%methylation and yellow circles indicate 0% methylation. The results from biological duplicate samples are shown. The bottom lane (Sequenom Methylation Analysis) shows the results of analysis of testis <t>DNA</t> after in vitro methylation using M. Sss1 methylase (New England Biolabs).
    Control Dna, supplied by Millipore, used in various techniques. Bioz Stars score: 93/100, based on 287 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Bio-Rad input dna
    ddPCR optimization based on existing SIV gag <t>DNA</t> real time qPCR assay and condition. (A) Direct migration of the qPCR SIV gag DNA assay in existing format onto <t>RainDance</t> ddPCR platform. Left: tissue DNA containing preamplified SIV DNA as template; Right: SIV DNA standard spike-in in buffer as template. (B) The effect of modifying MgCl 2 concentration on cluster separation. (C) Background issue in the SIV target detection region using existing assay’s master mix with optimized MgCl 2 concentration.
    Input Dna, supplied by Bio-Rad, used in various techniques. Bioz Stars score: 92/100, based on 780 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Illumina Inc input dna
    ddPCR optimization based on existing SIV gag <t>DNA</t> real time qPCR assay and condition. (A) Direct migration of the qPCR SIV gag DNA assay in existing format onto <t>RainDance</t> ddPCR platform. Left: tissue DNA containing preamplified SIV DNA as template; Right: SIV DNA standard spike-in in buffer as template. (B) The effect of modifying MgCl 2 concentration on cluster separation. (C) Background issue in the SIV target detection region using existing assay’s master mix with optimized MgCl 2 concentration.
    Input Dna, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 94/100, based on 2053 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Illumina Inc chip enriched dna
    ddPCR optimization based on existing SIV gag <t>DNA</t> real time qPCR assay and condition. (A) Direct migration of the qPCR SIV gag DNA assay in existing format onto <t>RainDance</t> ddPCR platform. Left: tissue DNA containing preamplified SIV DNA as template; Right: SIV DNA standard spike-in in buffer as template. (B) The effect of modifying MgCl 2 concentration on cluster separation. (C) Background issue in the SIV target detection region using existing assay’s master mix with optimized MgCl 2 concentration.
    Chip Enriched Dna, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 90/100, based on 169 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore salmon sperm dna protein a sepharose beads
    ddPCR optimization based on existing SIV gag <t>DNA</t> real time qPCR assay and condition. (A) Direct migration of the qPCR SIV gag DNA assay in existing format onto <t>RainDance</t> ddPCR platform. Left: tissue DNA containing preamplified SIV DNA as template; Right: SIV DNA standard spike-in in buffer as template. (B) The effect of modifying MgCl 2 concentration on cluster separation. (C) Background issue in the SIV target detection region using existing assay’s master mix with optimized MgCl 2 concentration.
    Salmon Sperm Dna Protein A Sepharose Beads, supplied by Millipore, used in various techniques. Bioz Stars score: 96/100, based on 13 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Upstate Biotechnology Inc salmon testis dna protein a agarose slurry
    ddPCR optimization based on existing SIV gag <t>DNA</t> real time qPCR assay and condition. (A) Direct migration of the qPCR SIV gag DNA assay in existing format onto <t>RainDance</t> ddPCR platform. Left: tissue DNA containing preamplified SIV DNA as template; Right: SIV DNA standard spike-in in buffer as template. (B) The effect of modifying MgCl 2 concentration on cluster separation. (C) Background issue in the SIV target detection region using existing assay’s master mix with optimized MgCl 2 concentration.
    Salmon Testis Dna Protein A Agarose Slurry, supplied by Upstate Biotechnology Inc, used in various techniques. Bioz Stars score: 85/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Illumina Inc chip seq dna sample prep kit
    ddPCR optimization based on existing SIV gag <t>DNA</t> real time qPCR assay and condition. (A) Direct migration of the qPCR SIV gag DNA assay in existing format onto <t>RainDance</t> ddPCR platform. Left: tissue DNA containing preamplified SIV DNA as template; Right: SIV DNA standard spike-in in buffer as template. (B) The effect of modifying MgCl 2 concentration on cluster separation. (C) Background issue in the SIV target detection region using existing assay’s master mix with optimized MgCl 2 concentration.
    Chip Seq Dna Sample Prep Kit, supplied by Illumina Inc, used in various techniques. Bioz Stars score: 89/100, based on 544 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Qiagen hotstartaq dna polymerase
    ddPCR optimization based on existing SIV gag <t>DNA</t> real time qPCR assay and condition. (A) Direct migration of the qPCR SIV gag DNA assay in existing format onto <t>RainDance</t> ddPCR platform. Left: tissue DNA containing preamplified SIV DNA as template; Right: SIV DNA standard spike-in in buffer as template. (B) The effect of modifying MgCl 2 concentration on cluster separation. (C) Background issue in the SIV target detection region using existing assay’s master mix with optimized MgCl 2 concentration.
    Hotstartaq Dna Polymerase, supplied by Qiagen, used in various techniques. Bioz Stars score: 99/100, based on 6724 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore total genomic dna input
    ddPCR optimization based on existing SIV gag <t>DNA</t> real time qPCR assay and condition. (A) Direct migration of the qPCR SIV gag DNA assay in existing format onto <t>RainDance</t> ddPCR platform. Left: tissue DNA containing preamplified SIV DNA as template; Right: SIV DNA standard spike-in in buffer as template. (B) The effect of modifying MgCl 2 concentration on cluster separation. (C) Background issue in the SIV target detection region using existing assay’s master mix with optimized MgCl 2 concentration.
    Total Genomic Dna Input, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Comparison of methylation data between MeDIP/NimbleGen Promoter + CpGi Array and Sequenom MassArray: Stk31 CpGi promoter region . The Stk31 gene region with a CpGi promoter region that is methylated in the liver, but not in ES or testis cells. The box indicates the position of the methylation peak in the liver that overlaps with the CpGi promoter region analyzed by Sequenom. In the epigram of Sequenom methylation analysis shown at the bottom panel, the blue circles indicate 100% methylation, green circles indicate around 50%methylation and yellow circles indicate 0% methylation. The results from biological duplicate samples are shown. The bottom lane (Sequenom Methylation Analysis) shows the results of analysis of testis DNA after in vitro methylation using M. Sss1 methylase (New England Biolabs).

    Journal: BMC Genomics

    Article Title: Genome-wide survey reveals dynamic widespread tissue-specific changes in DNA methylation during development

    doi: 10.1186/1471-2164-12-231

    Figure Lengend Snippet: Comparison of methylation data between MeDIP/NimbleGen Promoter + CpGi Array and Sequenom MassArray: Stk31 CpGi promoter region . The Stk31 gene region with a CpGi promoter region that is methylated in the liver, but not in ES or testis cells. The box indicates the position of the methylation peak in the liver that overlaps with the CpGi promoter region analyzed by Sequenom. In the epigram of Sequenom methylation analysis shown at the bottom panel, the blue circles indicate 100% methylation, green circles indicate around 50%methylation and yellow circles indicate 0% methylation. The results from biological duplicate samples are shown. The bottom lane (Sequenom Methylation Analysis) shows the results of analysis of testis DNA after in vitro methylation using M. Sss1 methylase (New England Biolabs).

    Article Snippet: After validating the enrichment of MeDIP DNA, MeDIP DNA and control input DNA were amplified by whole-genome amplification kit (Sigma Aldrich), followed by purification (QIAGEN Quick PCR Purification Kit) and then sent to NimbleGen for Microarray hybridization according to their standard protocol for the array.

    Techniques: Methylation, Methylated DNA Immunoprecipitation, In Vitro

    TgAP2X-5 depletion prevents TgAP2XI-5 binding to a number of promoters. ( A ) ChIP-on-chip data are represented as the log 2 ratio of the hybridization signal given by DNA immunoprecipitated over the signal given by the non-enriched input DNA. The log 2 ratio of each oligonucleotide present on the tiled microarray has been plotted at the respective genomic position. Genes above the horizontal axis are encoded the forward strand whereas those below are encoded the reverse strand. The scale of the Y -axis was kept identical for all representation to allow the comparison of signals. TgAP2XI-5 ChIP-chip assay is represented for the TgAP2XI-5 protein in the parental strain (RH ΔhxgprtΔku80 , green, negative control) or TgAP2XI-5 tagged strain (red, positive control) or TgAP2XI-5 tagged in the iKD TgAP2X-5 strain in presence of ATc (orange). The associated peaks identified by the mPeak algorithm are also provided. A portion of the Toxoplasma gondii chromosome III is represented. The boxed promoter presents a lower enrichment of TgAP2XI-5 in the iKD TgAP2X-5 strain in presence of ATc. ( B ) Representation of the ChIP-chip experiments for a portion of chromosome IV which encompasses the TgROP15 gene (yellow). Two repeats (R1 and R2) are presented for each ChIP experiment along with the mPeak identified peaks. ( C ) Representation of the ChIP-chip experiments for a portion of chromosome III which encompasses the TgROP24 gene (yellow). Two repeats (R1 and R2) are presented for each ChIP experiment along with the mPeak identified peaks. ( D ) Quantitative PCR was performed on the TgAP2XI-5 ChIP assays performed in the iKD TgAP2X-5/TgAP2XI-5 strain (orange), iKD TgAP2X-5 complemented/TgAP2XI-5 strain (yellow), TgAP2XI-5-tagged strain (red) and parental strain (negative control, green). Amplification was carried out on regions within the promoter of each of the six genes listed. The enrichment for corresponding ChIP DNA samples is presented as a percentage of INPUT for each target.

    Journal: Nucleic Acids Research

    Article Title: Cooperative binding of ApiAP2 transcription factors is crucial for the expression of virulence genes in Toxoplasma gondii

    doi: 10.1093/nar/gky373

    Figure Lengend Snippet: TgAP2X-5 depletion prevents TgAP2XI-5 binding to a number of promoters. ( A ) ChIP-on-chip data are represented as the log 2 ratio of the hybridization signal given by DNA immunoprecipitated over the signal given by the non-enriched input DNA. The log 2 ratio of each oligonucleotide present on the tiled microarray has been plotted at the respective genomic position. Genes above the horizontal axis are encoded the forward strand whereas those below are encoded the reverse strand. The scale of the Y -axis was kept identical for all representation to allow the comparison of signals. TgAP2XI-5 ChIP-chip assay is represented for the TgAP2XI-5 protein in the parental strain (RH ΔhxgprtΔku80 , green, negative control) or TgAP2XI-5 tagged strain (red, positive control) or TgAP2XI-5 tagged in the iKD TgAP2X-5 strain in presence of ATc (orange). The associated peaks identified by the mPeak algorithm are also provided. A portion of the Toxoplasma gondii chromosome III is represented. The boxed promoter presents a lower enrichment of TgAP2XI-5 in the iKD TgAP2X-5 strain in presence of ATc. ( B ) Representation of the ChIP-chip experiments for a portion of chromosome IV which encompasses the TgROP15 gene (yellow). Two repeats (R1 and R2) are presented for each ChIP experiment along with the mPeak identified peaks. ( C ) Representation of the ChIP-chip experiments for a portion of chromosome III which encompasses the TgROP24 gene (yellow). Two repeats (R1 and R2) are presented for each ChIP experiment along with the mPeak identified peaks. ( D ) Quantitative PCR was performed on the TgAP2XI-5 ChIP assays performed in the iKD TgAP2X-5/TgAP2XI-5 strain (orange), iKD TgAP2X-5 complemented/TgAP2XI-5 strain (yellow), TgAP2XI-5-tagged strain (red) and parental strain (negative control, green). Amplification was carried out on regions within the promoter of each of the six genes listed. The enrichment for corresponding ChIP DNA samples is presented as a percentage of INPUT for each target.

    Article Snippet: Amplification of immunoprecipitated DNA and 10 ng input DNA was performed using the GenomePlex® Complete Whole Amplification (WGA) kit (Sigma).

    Techniques: Binding Assay, Chromatin Immunoprecipitation, Hybridization, Immunoprecipitation, Microarray, Negative Control, Positive Control, Real-time Polymerase Chain Reaction, Amplification

    Assessment of the specificity of the ChIP protocol. A) Arx immunoprecipitation from N2a cells transfected with an Arx-expressing vector or a vector alone and using the Arx C-14 antibody from Santa Cruz. Detection using the anti-Arx-HD antibody revealed a major band at approximately 80 kD, which corresponds to the size of Arx protein. TE: total extracts, IP: immunoprecipitation, Ig: immunoglobulins. B) Enrichment of Ebf3 , Lmo1 and Shox2 promoter regions was assessed by qPCR using either Arx or PolII-immunoprecipitates or using no antibody and was compared to the total input. Gapdh was used as a positive control with DNA Pol II. Here, we show an example of the results obtained in a ChIP experiment with Arx C-14 antibody from Santa Cruz. The enrichment of Ebf3 , Lmo1 and Shox2 was checked similarly for each replicate experiment before DNA was applied to the microarrays.

    Journal: PLoS ONE

    Article Title: High-Throughput Analysis of Promoter Occupancy Reveals New Targets for Arx, a Gene Mutated in Mental Retardation and Interneuronopathies

    doi: 10.1371/journal.pone.0025181

    Figure Lengend Snippet: Assessment of the specificity of the ChIP protocol. A) Arx immunoprecipitation from N2a cells transfected with an Arx-expressing vector or a vector alone and using the Arx C-14 antibody from Santa Cruz. Detection using the anti-Arx-HD antibody revealed a major band at approximately 80 kD, which corresponds to the size of Arx protein. TE: total extracts, IP: immunoprecipitation, Ig: immunoglobulins. B) Enrichment of Ebf3 , Lmo1 and Shox2 promoter regions was assessed by qPCR using either Arx or PolII-immunoprecipitates or using no antibody and was compared to the total input. Gapdh was used as a positive control with DNA Pol II. Here, we show an example of the results obtained in a ChIP experiment with Arx C-14 antibody from Santa Cruz. The enrichment of Ebf3 , Lmo1 and Shox2 was checked similarly for each replicate experiment before DNA was applied to the microarrays.

    Article Snippet: DNA amplification, labelling and hybridization on microarrays Equivalent amounts of immunoprecipitated chromatin and total input DNA were amplified in parallel, using a random primer method with the GenomePlex Complete Whole Genome Amplification Kit (15 cycles), according to the manufacturer's instructions (Sigma).

    Techniques: Chromatin Immunoprecipitation, Immunoprecipitation, Transfection, Expressing, Plasmid Preparation, Real-time Polymerase Chain Reaction, Positive Control

    ChIP-chip results obtained from mouse embryonic brain. A) Graph representing log 2 probe intensities of Arx-immunoprecipitated DNA (IP) and input DNA obtained in a representative ChIP experiment. The red dots indicate the probes enriched in Arx-immunoprecipitates compared to total input DNA. B) Example of the enrichment profiles of Arx-bound promoter regions visualized by DNA analytics. The 3 lines represent data obtained from 3 independent ChIP-chip experiments and show the reproducibility between the 3 replicates. Contrarily to N2a cells, in which several continuous probes were often found enriched, only one or two probes were found enriched per gene. C) Venn diagram illustrating the overlap (black) between Arx-immunoprecipitated genes in transfected N2a cells (blue) and mouse embryonic brain (red), and the number of genes with at least 75% match to Arx-binding motif.

    Journal: PLoS ONE

    Article Title: High-Throughput Analysis of Promoter Occupancy Reveals New Targets for Arx, a Gene Mutated in Mental Retardation and Interneuronopathies

    doi: 10.1371/journal.pone.0025181

    Figure Lengend Snippet: ChIP-chip results obtained from mouse embryonic brain. A) Graph representing log 2 probe intensities of Arx-immunoprecipitated DNA (IP) and input DNA obtained in a representative ChIP experiment. The red dots indicate the probes enriched in Arx-immunoprecipitates compared to total input DNA. B) Example of the enrichment profiles of Arx-bound promoter regions visualized by DNA analytics. The 3 lines represent data obtained from 3 independent ChIP-chip experiments and show the reproducibility between the 3 replicates. Contrarily to N2a cells, in which several continuous probes were often found enriched, only one or two probes were found enriched per gene. C) Venn diagram illustrating the overlap (black) between Arx-immunoprecipitated genes in transfected N2a cells (blue) and mouse embryonic brain (red), and the number of genes with at least 75% match to Arx-binding motif.

    Article Snippet: DNA amplification, labelling and hybridization on microarrays Equivalent amounts of immunoprecipitated chromatin and total input DNA were amplified in parallel, using a random primer method with the GenomePlex Complete Whole Genome Amplification Kit (15 cycles), according to the manufacturer's instructions (Sigma).

    Techniques: Chromatin Immunoprecipitation, Immunoprecipitation, Transfection, Binding Assay

    ChIP-chip results obtained from Arx-transfected N2a cells. A) Graph representing log 2 probe intensities of Arx-immunoprecipitated DNA (IP) and input DNA obtained in a representative ChIP experiment. The red dots indicate the probes enriched in Arx-immunoprecipitates compared to total input DNA. B) Examples of the enrichment profiles of Arx-bound promoter regions visualized by DNA analytics software. The 3 lines represent data obtained from 3 independent ChIP-chip experiments and show the reproducibility between the 3 replicates. C) The resulting weighted matrix discovered through the MDModule analysis (top) appears to be similar to the motif identified by Berger et al. [27] (bottom). D) Frequency distribution of scores. The TAATTA motif identified by MDModule was significantly more present in ChIP-enriched sequences (red curve) by comparison to negative control sequences (blue curve).

    Journal: PLoS ONE

    Article Title: High-Throughput Analysis of Promoter Occupancy Reveals New Targets for Arx, a Gene Mutated in Mental Retardation and Interneuronopathies

    doi: 10.1371/journal.pone.0025181

    Figure Lengend Snippet: ChIP-chip results obtained from Arx-transfected N2a cells. A) Graph representing log 2 probe intensities of Arx-immunoprecipitated DNA (IP) and input DNA obtained in a representative ChIP experiment. The red dots indicate the probes enriched in Arx-immunoprecipitates compared to total input DNA. B) Examples of the enrichment profiles of Arx-bound promoter regions visualized by DNA analytics software. The 3 lines represent data obtained from 3 independent ChIP-chip experiments and show the reproducibility between the 3 replicates. C) The resulting weighted matrix discovered through the MDModule analysis (top) appears to be similar to the motif identified by Berger et al. [27] (bottom). D) Frequency distribution of scores. The TAATTA motif identified by MDModule was significantly more present in ChIP-enriched sequences (red curve) by comparison to negative control sequences (blue curve).

    Article Snippet: DNA amplification, labelling and hybridization on microarrays Equivalent amounts of immunoprecipitated chromatin and total input DNA were amplified in parallel, using a random primer method with the GenomePlex Complete Whole Genome Amplification Kit (15 cycles), according to the manufacturer's instructions (Sigma).

    Techniques: Chromatin Immunoprecipitation, Transfection, Immunoprecipitation, Software, Negative Control

    Confirmation of Arx binding to candidate promoter regions. A) Arx-immunoprecipitated DNA was compared to input DNA by ChIP/QFM-PCR to determine ChIP enrichment of 21 putative target genes. No enrichment occurred for the Vapb promoter which was consistently negative in all ChIP experiments. Bar heights represent log 10 enrichment of the signal obtained for Arx-immunoprecipitated DNA versus input DNA for each promoter using site-specific primers. The arrows above the bars indicate sequences in which the previously defined Arx-binding motif was found. B) Confirmation of Arx binding to 5 different promoter sequences identified by ChIP using a luciferase reporter gene assay. Firefly luciferase data were normalized to Renilla luciferase expression and data are presented as the percentage of transcriptional activity compared to the vector control. Arx regulation was confirmed for Lmo1 , Lmo3 , Sh3tc2 , Calb2 and Cdh2 promoter regions as transcriptional activity was repressed in the presence of Arx by comparison to the control transfection, whereas it had no effect on the plasmid alone pGL4.23. Error bars indicate SEM.

    Journal: PLoS ONE

    Article Title: High-Throughput Analysis of Promoter Occupancy Reveals New Targets for Arx, a Gene Mutated in Mental Retardation and Interneuronopathies

    doi: 10.1371/journal.pone.0025181

    Figure Lengend Snippet: Confirmation of Arx binding to candidate promoter regions. A) Arx-immunoprecipitated DNA was compared to input DNA by ChIP/QFM-PCR to determine ChIP enrichment of 21 putative target genes. No enrichment occurred for the Vapb promoter which was consistently negative in all ChIP experiments. Bar heights represent log 10 enrichment of the signal obtained for Arx-immunoprecipitated DNA versus input DNA for each promoter using site-specific primers. The arrows above the bars indicate sequences in which the previously defined Arx-binding motif was found. B) Confirmation of Arx binding to 5 different promoter sequences identified by ChIP using a luciferase reporter gene assay. Firefly luciferase data were normalized to Renilla luciferase expression and data are presented as the percentage of transcriptional activity compared to the vector control. Arx regulation was confirmed for Lmo1 , Lmo3 , Sh3tc2 , Calb2 and Cdh2 promoter regions as transcriptional activity was repressed in the presence of Arx by comparison to the control transfection, whereas it had no effect on the plasmid alone pGL4.23. Error bars indicate SEM.

    Article Snippet: DNA amplification, labelling and hybridization on microarrays Equivalent amounts of immunoprecipitated chromatin and total input DNA were amplified in parallel, using a random primer method with the GenomePlex Complete Whole Genome Amplification Kit (15 cycles), according to the manufacturer's instructions (Sigma).

    Techniques: Binding Assay, Immunoprecipitation, Chromatin Immunoprecipitation, Polymerase Chain Reaction, Luciferase, Reporter Gene Assay, Expressing, Activity Assay, Plasmid Preparation, Transfection

    ddPCR optimization based on existing SIV gag DNA real time qPCR assay and condition. (A) Direct migration of the qPCR SIV gag DNA assay in existing format onto RainDance ddPCR platform. Left: tissue DNA containing preamplified SIV DNA as template; Right: SIV DNA standard spike-in in buffer as template. (B) The effect of modifying MgCl 2 concentration on cluster separation. (C) Background issue in the SIV target detection region using existing assay’s master mix with optimized MgCl 2 concentration.

    Journal: PLoS ONE

    Article Title: Maximizing viral detection with SIV droplet digital PCR (ddPCR) assays

    doi: 10.1371/journal.pone.0233085

    Figure Lengend Snippet: ddPCR optimization based on existing SIV gag DNA real time qPCR assay and condition. (A) Direct migration of the qPCR SIV gag DNA assay in existing format onto RainDance ddPCR platform. Left: tissue DNA containing preamplified SIV DNA as template; Right: SIV DNA standard spike-in in buffer as template. (B) The effect of modifying MgCl 2 concentration on cluster separation. (C) Background issue in the SIV target detection region using existing assay’s master mix with optimized MgCl 2 concentration.

    Article Snippet: In conclusion, combining two ddPCR assays for SIV nucleic acids detection with the RainDance ddPCR platform can enable a large amount of input DNA to be analyzed per reaction, and can overcome severe RNA inhibition when combined with suitable reverse transcription enzyme(s).

    Techniques: Real-time Polymerase Chain Reaction, Migration, Concentration Assay

    Comparison between ddPCR and qPCR quantitation results in low viral DNA range

    Journal: PLoS ONE

    Article Title: Maximizing viral detection with SIV droplet digital PCR (ddPCR) assays

    doi: 10.1371/journal.pone.0233085

    Figure Lengend Snippet: Comparison between ddPCR and qPCR quantitation results in low viral DNA range

    Article Snippet: In conclusion, combining two ddPCR assays for SIV nucleic acids detection with the RainDance ddPCR platform can enable a large amount of input DNA to be analyzed per reaction, and can overcome severe RNA inhibition when combined with suitable reverse transcription enzyme(s).

    Techniques: Real-time Polymerase Chain Reaction, Quantitation Assay

    SIV ddPCR DNA assay characterization. (A) Sample DNA input tolerance at the droplet formation step. Droplet integrity was monitored by examining a portion of the droplets in each lane as they moved through the Source instrument during dropletization. In addition, total droplet number for each input level after dropletization (retrieved from the “RainDrop Run Completion” screen) served as another indicator of sample DNA input tolerance. DNA sample used was duodenum DNA from Rhesus macaque 313–08. (B) Total number of droplets generated during dropletization remains constant in the range of tissue DNA amount tested (1 million to 8 million cell equivalent input). Total droplet number for each sample after dropletization serves as an additional indicator of sample DNA input tolerance. Note that for each level of DNA input, only a fraction (~1.6%) of the droplets were counted for QC purpose by the Source machine. (C) Estimation of the limit of detection (LoD) of the ddPCR assay based on the Digital MIQE Guidelines [ 7 ]. According to the guidelines, when running costs preclude optimization using ddPCR, qPCR can be used to determine certain assay parameters. (D) Performance of the SIV ddPCR assay in TaqMan genotyping mastermix in qPCR format. SIV gag DNA standard was diluted with buffer diluent. The standards were assayed as described in Materials and Methods in the following replicate format: 1 million down to 100 copies input per reaction: each in triplicates; 50, 20, 10, 7 and 5 copies input per reaction: each in 10 replicates. The data were plotted and analyzed according to the routine analyses provided in the software package with the ABI 7500 SDS instrument. (E) Instead of measuring 60 ddPCR replicates to obtain 95% confidence, we obtained an approximate estimation of the LoD using a lower number of ddPCR reaction replicates and required the false negative rate to be below 5% (i.e. all 10 replicates have to be positive).

    Journal: PLoS ONE

    Article Title: Maximizing viral detection with SIV droplet digital PCR (ddPCR) assays

    doi: 10.1371/journal.pone.0233085

    Figure Lengend Snippet: SIV ddPCR DNA assay characterization. (A) Sample DNA input tolerance at the droplet formation step. Droplet integrity was monitored by examining a portion of the droplets in each lane as they moved through the Source instrument during dropletization. In addition, total droplet number for each input level after dropletization (retrieved from the “RainDrop Run Completion” screen) served as another indicator of sample DNA input tolerance. DNA sample used was duodenum DNA from Rhesus macaque 313–08. (B) Total number of droplets generated during dropletization remains constant in the range of tissue DNA amount tested (1 million to 8 million cell equivalent input). Total droplet number for each sample after dropletization serves as an additional indicator of sample DNA input tolerance. Note that for each level of DNA input, only a fraction (~1.6%) of the droplets were counted for QC purpose by the Source machine. (C) Estimation of the limit of detection (LoD) of the ddPCR assay based on the Digital MIQE Guidelines [ 7 ]. According to the guidelines, when running costs preclude optimization using ddPCR, qPCR can be used to determine certain assay parameters. (D) Performance of the SIV ddPCR assay in TaqMan genotyping mastermix in qPCR format. SIV gag DNA standard was diluted with buffer diluent. The standards were assayed as described in Materials and Methods in the following replicate format: 1 million down to 100 copies input per reaction: each in triplicates; 50, 20, 10, 7 and 5 copies input per reaction: each in 10 replicates. The data were plotted and analyzed according to the routine analyses provided in the software package with the ABI 7500 SDS instrument. (E) Instead of measuring 60 ddPCR replicates to obtain 95% confidence, we obtained an approximate estimation of the LoD using a lower number of ddPCR reaction replicates and required the false negative rate to be below 5% (i.e. all 10 replicates have to be positive).

    Article Snippet: In conclusion, combining two ddPCR assays for SIV nucleic acids detection with the RainDance ddPCR platform can enable a large amount of input DNA to be analyzed per reaction, and can overcome severe RNA inhibition when combined with suitable reverse transcription enzyme(s).

    Techniques: Generated, Real-time Polymerase Chain Reaction, Software

    ddPCR and qPCR comparison. (A) Sample inhibition comparison between ddPCR and qPCR. An ovarian DNA sample (Rhesus macaque 311–08) was subjected to nested (i.e. preamplified) qPCR analysis or direct RainDance (“RD” in the table header) ddPCR analysis for SIV DNA viral load. At 3.7 million cell input per reaction (at the preamp step), SIV qPCR signal is inhibited by 99%, while at 4 M cell input per direct ddPCR reaction, SIV signal is not inhibited. Note that in the qPCR graph, the data point (32.6 ± 5.4) on the left (0.37 million cell input per reaction) was derived from a standard curve, while the data point (4.0) on the right (3.7 million cell input per reaction) was derived from Poisson statistics (i.e. DNA copies derived from the positive well rate among the replicate reactions). (B) Quantification of Rhesus macaque necropsy tissue DNA samples (from Rhesus macaque 27882) (including an uninfected negative control sample) for SIV DNA using ddPCR. (C) Comparison between ddPCR and nested qPCR analysis results for the necropsy tissue samples in (B). RD, RainDance. S.D., standard deviation.

    Journal: PLoS ONE

    Article Title: Maximizing viral detection with SIV droplet digital PCR (ddPCR) assays

    doi: 10.1371/journal.pone.0233085

    Figure Lengend Snippet: ddPCR and qPCR comparison. (A) Sample inhibition comparison between ddPCR and qPCR. An ovarian DNA sample (Rhesus macaque 311–08) was subjected to nested (i.e. preamplified) qPCR analysis or direct RainDance (“RD” in the table header) ddPCR analysis for SIV DNA viral load. At 3.7 million cell input per reaction (at the preamp step), SIV qPCR signal is inhibited by 99%, while at 4 M cell input per direct ddPCR reaction, SIV signal is not inhibited. Note that in the qPCR graph, the data point (32.6 ± 5.4) on the left (0.37 million cell input per reaction) was derived from a standard curve, while the data point (4.0) on the right (3.7 million cell input per reaction) was derived from Poisson statistics (i.e. DNA copies derived from the positive well rate among the replicate reactions). (B) Quantification of Rhesus macaque necropsy tissue DNA samples (from Rhesus macaque 27882) (including an uninfected negative control sample) for SIV DNA using ddPCR. (C) Comparison between ddPCR and nested qPCR analysis results for the necropsy tissue samples in (B). RD, RainDance. S.D., standard deviation.

    Article Snippet: In conclusion, combining two ddPCR assays for SIV nucleic acids detection with the RainDance ddPCR platform can enable a large amount of input DNA to be analyzed per reaction, and can overcome severe RNA inhibition when combined with suitable reverse transcription enzyme(s).

    Techniques: Real-time Polymerase Chain Reaction, Inhibition, Derivative Assay, Negative Control, Standard Deviation

    Identifying an optimal ddPCR assay format and condition. Performance of an MGB probe-based SIV gag ddPCR assay using TaqMan genotyping mastermix on (A) SIV (on CCR5 background) spike-in templates and (B) ovary tissue DNA from a Rhesus macaque (311–08) infected with SIVmac239. (C) A summary of all other tested conditions (probe and mastermix) and the observed issue(s) associated with each condition.

    Journal: PLoS ONE

    Article Title: Maximizing viral detection with SIV droplet digital PCR (ddPCR) assays

    doi: 10.1371/journal.pone.0233085

    Figure Lengend Snippet: Identifying an optimal ddPCR assay format and condition. Performance of an MGB probe-based SIV gag ddPCR assay using TaqMan genotyping mastermix on (A) SIV (on CCR5 background) spike-in templates and (B) ovary tissue DNA from a Rhesus macaque (311–08) infected with SIVmac239. (C) A summary of all other tested conditions (probe and mastermix) and the observed issue(s) associated with each condition.

    Article Snippet: In conclusion, combining two ddPCR assays for SIV nucleic acids detection with the RainDance ddPCR platform can enable a large amount of input DNA to be analyzed per reaction, and can overcome severe RNA inhibition when combined with suitable reverse transcription enzyme(s).

    Techniques: Infection

    SIV ddPCR DNA assay performance. (A) SIV ddPCR DNA assay detection of low single digit level SIV DNA input and linear dynamic range of the SIV ddPCR DNA assay. Different amount of SIV DNA template was spiked in 1 million Rhesus macaque PBMC equivalent of genomic DNA background each. SIV DNA amount was quantified by direct ddPCR. (B) Quantification data of Fig 5A.

    Journal: PLoS ONE

    Article Title: Maximizing viral detection with SIV droplet digital PCR (ddPCR) assays

    doi: 10.1371/journal.pone.0233085

    Figure Lengend Snippet: SIV ddPCR DNA assay performance. (A) SIV ddPCR DNA assay detection of low single digit level SIV DNA input and linear dynamic range of the SIV ddPCR DNA assay. Different amount of SIV DNA template was spiked in 1 million Rhesus macaque PBMC equivalent of genomic DNA background each. SIV DNA amount was quantified by direct ddPCR. (B) Quantification data of Fig 5A.

    Article Snippet: In conclusion, combining two ddPCR assays for SIV nucleic acids detection with the RainDance ddPCR platform can enable a large amount of input DNA to be analyzed per reaction, and can overcome severe RNA inhibition when combined with suitable reverse transcription enzyme(s).

    Techniques: