Extensive chromatin fragmentation improves enrichment of protein binding sites in chromatin immunoprecipitation experiments
Open Access
- 2 September 2008
- journal article
- research article
- Published by Oxford University Press (OUP) in Nucleic Acids Research
- Vol. 36 (19), e125
- https://doi.org/10.1093/nar/gkn535
Abstract
Extensive sonication of formaldehyde-crosslinked chromatin can generate DNA fragments averaging 200 bp in length (range 75–300 bp). Fragmentation is largely random with respect to genomic region and nucleosome position. ChIP experiments employing such extensively fragmented samples show 2- to 4-fold increased enrichment of protein binding sites over control genomic regions, when compared to samples sonicated to a more conventional size range (300–500 bp). The basis of improved fold enrichments is that immunoprecipitation of protein-bound regions is unaffected by fragment size, whereas immunoprecipitation of control genomic regions decreases progressively along with reduced fragment size due to fewer nonspecific binding sites. The use of extensively sonicated samples improves mapping of protein binding sites, and it extends the dynamic range for quantitative measurements of histone density. We show that many yeast promoter regions are virtually devoid of histones.Keywords
This publication has 19 references indexed in Scilit:
- Genome-wide profiles of STAT1 DNA association using chromatin immunoprecipitation and massively parallel sequencingNature Methods, 2007
- Genome-Wide Mapping of in Vivo Protein-DNA InteractionsScience, 2007
- Genome-Scale Identification of Nucleosome Positions in S. cerevisiaeScience, 2005
- Intrinsic Histone-DNA Interactions and Low Nucleosome Density Are Important for Preferential Accessibility of Promoter Regions in YeastMolecular Cell, 2005
- Chromatin Immunoprecipitation for Determining the Association of Proteins with Specific Genomic Sequences In VivoCurrent Protocols in Molecular Biology, 2005
- Evidence for Eviction and Rapid Deposition of Histones upon Transcriptional Elongation by RNA Polymerase IIMolecular and Cellular Biology, 2004
- Defining the CREB RegulonCell, 2004
- Evidence for nucleosome depletion at active regulatory regions genome-wideNature Genetics, 2004
- Application of the split-ubiquitin membrane yeast two-hybrid system to investigate membrane protein interactionsMethods, 2004
- Targeted Recruitment of Set1 Histone Methylase by Elongating Pol II Provides a Localized Mark and Memory of Recent Transcriptional ActivityMolecular Cell, 2003