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Calibrating ChIP-seq with nucleosomal internal standards to measure histone modification density genome-wide

ABSTRACT: Calibrating ChIP-seq with nucleosomal internal standards to measure histone modification density genome-wide

PROVIDER: PRJNA258581 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR1555110_1.fastq.gz	Fastqsanger.gz
	SRR1555110_2.fastq.gz	Fastqsanger.gz
	SRR1555111_1.fastq.gz	Fastqsanger.gz
	SRR1555111_2.fastq.gz	Fastqsanger.gz
	SRR1555112_1.fastq.gz	Fastqsanger.gz

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Calibrating ChIP-seq with nucleosomal internal standards to measure histone modification density genome-wide

Project description:Despite serving as a central experimental technique in epigenetics research, chromatin immunoprecipitation (ChIP) suffers from several serious drawbacks: it is a relative measurement untethered to any external scale that obviates fair comparison amongst experiments; it employs antibody reagents that have differing affinity and specificity for target epitopes, which are in turn variable in abundance; and it is frequently not reproducible. To address these problems, we developed internal standard calibrated ChIP (ICeChIP), a method of spiking a native chromatin sample with nucleosomes reconstituted from recombinant and semisynthetic histones on barcoded DNA prior to immunoprecipitation. ICeChIP measures local histone modification densities on a biologically meaningful scale, enabling unbiased trans-experimental comparisons and revealing a correlation between the apparent symmetry of H3K4me3 in promoter nucleosomes and gene expression. Direct in situ assessment of immunoprecipitation accommodates for a number of experimental pitfalls, and provides a critical examination of untested assumptions inherent in conventional ChIP.

2015-05-07 | GSE60378 | GEO

Calibrating ChIP-seq with nucleosomal internal standards to measure histone modification density genome-wide

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Calibrating ChIP-Seq with Nucleosomal Internal Standards to Measure Histone Modification Density Genome Wide.

Project description:Chromatin immunoprecipitation (ChIP) serves as a central experimental technique in epigenetics research, yet there are serious drawbacks: it is a relative measurement, which untethered to any external scale obscures fair comparison among experiments; it employs antibody reagents that have differing affinities and specificities for target epitopes that vary in abundance; and it is frequently not reproducible. To address these problems, we developed Internal Standard Calibrated ChIP (ICeChIP), wherein a native chromatin sample is spiked with nucleosomes reconstituted from recombinant and semisynthetic histones on barcoded DNA prior to immunoprecipitation. ICeChIP measures local histone modification densities on a biologically meaningful scale, enabling unbiased trans-experimental comparisons, and reveals unique insight into the nature of bivalent domains. This technology provides in situ assessment of the immunoprecipitation step, accommodating for many experimental pitfalls as well as providing a critical examination of untested assumptions inherent to conventional ChIP.

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