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Efficient double fragmentation ChIP-seq provides nucleotide resolution protein-DNA binding profiles

Mokry, M. and Hatzis, P. and Bruijn de, E. and Koster, J. and Versteeg, R. and Schuijers, J. and Wetering van de, M.L. and Guryev, V. and Clevers, H. and Cuppen, E. (2010) Efficient double fragmentation ChIP-seq provides nucleotide resolution protein-DNA binding profiles. PLoS One, 5, e15092-. ISSN 1932-6203.

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Immunoprecipitated crosslinked protein-DNA fragments typically range in size from several hundred to several thousand base pairs, with a significant part of chromatin being much longer than the optimal length for next-generation sequencing (NGS) procedures. Because these larger fragments may be non-random and represent relevant biology that may otherwise be missed, but also because they represent a significant fraction of the immunoprecipitated material, we designed a double-fragmentation ChIP-seq procedure. After conventional crosslinking and immunoprecipitation, chromatin is de-crosslinked and sheared a second time to concentrate fragments in the optimal size range for NGS. Besides the benefits of increased chromatin yields, the procedure also eliminates a laborious size-selection step. We show that the double-fragmentation ChIP-seq approach allows for the generation of biologically relevant genome-wide protein-DNA binding profiles from sub-nanogram amounts of TCF7L2/TCF4, TBP and H3K4me3 immunoprecipitated material. Although optimized for the AB/SOLiD platform, the same approach may be applied to other platforms.

Item Type:Article
Institutes:Hubrecht Instituut
ID Code:9525
Deposited On:11 Feb 2011 01:00
Last Modified:07 Sep 2011 17:01

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