Chromatin Immunoprecipitation (ChIP)

Abstract

Chromatin immunoprecipitation or ChIP is an excellent method of investigation of the specific protein interaction and its altered forms with DNA region. These interactions have a significant role in various cellular processes such as replication, transcription, DNA damage repair, genome stability, gene regulation and segregation at mitosis. This technique is therefore giving us power to study a variety of cellular mechanisms inside the cell in terms of protein-DNA interaction. As the name Chromatin immunoprecipitation suggests this method utilizes chromatin preparation from cells to selectively immune-precipitate the protein of interest to identify DNA sequence associated with it. Chromatin is an organized structure of eukaryotic DNA which contains double-stranded DNA wrapped around nucleosomes. ChIP has been extensively used to depict transcription factors, variants of histone, chromatin modifying enzymes, post-translational modification of histone on the genome. In the classical ChIP method, protein and DNA is irreversibly cross-linked by UV exposure followed by immunoprecipitation with specific antibodies, protein-DNA complex is then purified, treated with proteases and then analysis is done by the method of Southern blot or dot blot using a radio-labelled probe derived from the cloned DNA fragment of interest. Further, it was modified by using formaldehyde for reversible cross-linking of protein-DNA complex and polymerase chain reaction for the detection of fragments of precipitated DNA. ChIP is a cumbersome procedure to perform and present many limitations, for example it requires many cells. Therefore, many modifications and variations, have also developed with the time which enables us to simplify the procedure and widen its range of applications. This chapter provides a brief method for Chromatin immunoprecipitation (ChIP) and its applications.