Unknown,Transcriptomics,Genomics,Proteomics

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Limitations and possibilities of low cell number ChIP-seq


ABSTRACT: ChIP-seq experiments using low numbers of input cells, scaled down to the point where data quality is unnacceptably compromised, reveals limits of the technique. Two-part ChIPseq study using native chromatin (non-crosslinked) generated with MNase from human CD4+ cells. Part 1: Previously published protocol (Barski et al, 2007, Cell 129:823, hereafter called "Benchmark") used with 2x10e7 cells / ChIP with H3K4me3 antibody, compared to modified method ("new") with decreasing input cell numbers spanning 1000-fold range from 2x10e7 cells / IP to 2x10e4 cells/ IP. Part 2: reproducibility of new method studied using triplicate samples at lowest two cell numbers tested: 1x10e5 and 2x10e4 / ChIP, using H3K4me3 and H3K27me3 antisera.

ORGANISM(S): Homo sapiens

SUBMITTER: Gregor Gilfillan 

PROVIDER: E-GEOD-42147 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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<h4>Background</h4>Chromatin immunoprecipitation coupled with high-throughput DNA sequencing (ChIP-seq) offers high resolution, genome-wide analysis of DNA-protein interactions. However, current standard methods require abundant starting material in the range of 1-20 million cells per immunoprecipitation, and remain a bottleneck to the acquisition of biologically relevant epigenetic data. Using a ChIP-seq protocol optimised for low cell numbers (down to 100,000 cells/IP), we examined the perform  ...[more]

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