Examining DNA–Protein Interactions with Genome-Wide Chromatin Immunoprecipitation Analysis
Understanding the mechanisms by which genomic information is hierarchically organized and used by different cell and tissue types under different physiological conditions requires the detailed analysis of the chromatin structure and nuclear factor distribution throughout the entire genome. Chromatin organization and histone modification patterns ultimately define cell identity, and the occurrence of aberrant changes in chromatin result in malfunction and disease. The strategy of systematically mapping the distribution of histone modifications, nucleosome positioning, and nuclear factor occupancy requires key methods for mapping DNA-protein interactions at the genome-wide level.
The use of chromatin immunoprecipitation (ChIP) assays, where an immunoprecipitating antibody against a particular factor or histone modification is used to enrich chromatin fractions in the sequences to which the protein is bound, has been very useful for defining the histone modification status and nuclear factor association at specific sites. More recently, the combination of ChIP assays with hybridization of microarrays (ChIP-chip) or with next generation massively parallel sequencing (ChIP-seq) has become an important strategy for the acquisition of this type of information at the genome-wide level. In this chapter, we present a critical overview of the considerations necessary for the design and execution of successful genome-wide ChIP experiments.
KeywordsSodium Dodecyl Sulfate Histone Modification ChIP Assay Whole Genome Amplification Chromatin Feature
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