cChIP-seq: a robust small-scale method for investigation of histone modifications View Full Text


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Article Info

DATE

2015-12-21

AUTHORS

Cristina Valensisi, Jo Ling Liao, Colin Andrus, Stephanie L. Battle, R. David Hawkins

ABSTRACT

BackgroundChIP-seq is highly utilized for mapping histone modifications that are informative about gene regulation and genome annotations. For example, applying ChIP-seq to histone modifications such as H3K4me1 has facilitated generating epigenomic maps of putative enhancers. This powerful technology, however, is limited in its application by the large number of cells required. ChIP-seq involves extensive manipulation of sample material and multiple reactions with limited quality control at each step, therefore, scaling down the number of cells required has proven challenging. Recently, several methods have been proposed to overcome this limit but most of these methods require extensive optimization to tailor the protocol to the specific antibody used or number of cells being profiled.ResultsHere we describe a robust, yet facile method, which we named carrier ChIP-seq (cChIP-seq), for use on limited cell amounts. cChIP-seq employs a DNA-free histone carrier in order to maintain the working ChIP reaction scale, removing the need to tailor reactions to specific amounts of cells or histone modifications to be assayed. We have applied our method to three different histone modifications, H3K4me3, H3K4me1 and H3K27me3 in the K562 cell line, and H3K4me1 in H1 hESCs. We successfully obtained epigenomic maps for these histone modifications starting with as few as 10,000 cells. We compared cChIP-seq data to data generated as part of the ENCODE project. ENCODE data are the reference standard in the field and have been generated starting from tens of million of cells. Our results show that cChIP-seq successfully recapitulates bulk data. Furthermore, we showed that the differences observed between small-scale ChIP-seq data and ENCODE data are largely to be due to lab-to-lab variability rather than operating on a reduced scale.ConclusionsData generated using cChIP-seq are equivalent to reference epigenomic maps from three orders of magnitude more cells. Our method offers a robust and straightforward approach to scale down ChIP-seq to as low as 10,000 cells. The underlying principle of our strategy makes it suitable for being applied to a vast range of chromatin modifications without requiring expensive optimization. Furthermore, our strategy of a DNA-free carrier can be adapted to most ChIP-seq protocols. More... »

PAGES

1083

References to SciGraph publications

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  • 2015-02-05. Amplification of pico-scale DNA mediated by bacterial carrier DNA for small-cell-number transcription factor ChIP-seq in BMC GENOMICS
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  • 2012-10-23. ChIP–seq and beyond: new and improved methodologies to detect and characterize protein–DNA interactions in NATURE REVIEWS GENETICS
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  • 2010-07-11. Genome-wide chromatin maps derived from limited numbers of hematopoietic progenitors in NATURE METHODS
  • 2015-02-18. Integrative analysis of 111 reference human epigenomes in NATURE
  • 2013-10-06. Transposition of native chromatin for fast and sensitive epigenomic profiling of open chromatin, DNA-binding proteins and nucleosome position in NATURE METHODS
  • 2009-09-08. ChIP–seq: advantages and challenges of a maturing technology in NATURE REVIEWS GENETICS
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