Chromatin IP
The Nexus of Genomics & Proteomics
Chromatin Immunoprecipitation: the Nexus of Genomics and Proteomics
Critical to understanding the biological function of a particular histone modification or chromosomal protein is the identification of its distribution throughout the genome and how that distribution is altered as part of specific biological processes. Currently, the chromatin immunoprecipitation (ChIP) technique is the most precise method to identify specific proteins associated with a region of the genome or, conversely, identification of regions of the genome associated with specific proteins. The proteins in question might be modified isoforms of histones or non-histone chromosomal proteins. Thus, one can use ChIP to quantitate the relative amount of histone H3 acetylation, for example, that is associated with a specific gene promoter under various conditions that alter expression of the gene. However, much of the recent usage of ChIP has been for identifying and analyzing transcription factor binding sites.
Schematic representation of the chromatin immunoprecipitation (ChIP) technique. First, the proteins are cross-linked to DNA with formaldehyde, and then the chromatin is sheared to a manageable size by sonication. Specific proteins are immunoprecipitated with antibodies, also bringing down the DNA to which the protein is cross-linked. The cross-links are reversed, the DNA purified, and the sample is interrogated for the enrichment of specific DNA sequences. The detection step can be performed most accurately by quantitative real-time PCR, but the usage of microarrays in this step is increasing. |
New techniques that combine the ChIP technique with microarray analysis facilitate the determination of genome-wide distribution patterns for chromosomal proteins and histone modifications. Using this combined technique, researchers can ask sweeping questions regarding the cellular functions of histone modifications and transcription factors and place these in the larger context of regulatory network interactions.
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