Title: Chromatin Immunoprecipitation in the Analysis of Large Chromatin Domains Across Murine Antigen Receptor Loci
Abstract: This chapter provides a full, detailed description of chromatin immunoprecipitation (ChIP) method, including the subsequent analysis of the immunoprecipitated DNA. ChIP is a powerful technique used to analyze protein or DNA interactions in a native chromatin context. In conjunction with previously favored procedures such as nuclease sensitivity mapping, ChIP also provides an additional tool to search for specific DNA regulatory elements. The chapter presents a ChIP protocol used to identify the histone modification patterns at murine immunoglobulin heavy chain (IgH) locus. Data generated from ChIP experiments provides valuable insight into how specific gene segments within a locus are made accessible to the recombination machinery while simultaneously restricting access to other gene segments during lymphocyte development. The ChIP method involves three basic steps, namely— (1) In vivo formaldehyde crosslinking of intact cells, (2) Micrococcal nuclease (MNase) digestion and sonication of the chromatin to generate a pool of small chromatin fragments, and (3) Selective immunoprecipitation (IP) of protein or DNA complexes from the chromatin pool with specific antibodies. The ChIP method described in this chapter provides another versatile tool, in addition to DNaseI hypersensitivity mapping, which can be used in a number of unique cell types to identify regulatory elements, such as promoters, enhancers, or boundary elements as well as define the histone modification pattern across chromatin sub-domains within a large, complex genomic locus.
Publication Year: 2003
Publication Date: 2003-01-01
Language: en
Type: review
Indexed In: ['crossref', 'pubmed']
Access and Citation
Cited By Count: 25
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