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

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Abstract

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 decrosslinked 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 doublefragmentation 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. © 2010 Mokry et al.

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Mokry, M., Hatzis, P., de Bruijn, E., Koster, J., Versteeg, R., Schuijers, J., … Cuppen, E. (2010). Efficient double fragmentation ChiP-seq provides nucleotide resolution protein-DNA binding profiles. PLoS ONE, 5(11). https://doi.org/10.1371/journal.pone.0015092

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