Histone modifications and transcription factor binding on chromatin ChIP-PCR assays.
Chromatin, the eukaryotic template of genetic information, is subject to a diverse array of posttranslational modifications that largely impinge on the N-termini of histones, such as acetylation, methylation, phosphorylation, and ubiquitination. Distinct histone modifications generate synergistic or antagonistic interaction affinities for novhistone proteins, which in turn dictate dynamic transitions between transcriptionally active or silent states of chromatin. Besides transcription, numerous biological processes, including DNA replication, DNA repair, and recombination, are regulated by chromatin-associated factors. The chromatin immunoprecipitation (ChIP) technique provides us with an exquisite tool to investigate the interplay between the structural or regulatory proteins and DNA and its role in regulating diverse cellular processes in vivo by formaldehyde crosslinking of proteins to proteins and proteins to DNA, followed by immunoprecipitation of the fixed material and detection of the associated DNA. Here we illustrate the overall experimental procedure by taking ChIP analysis of the human telomerase reverse transcriptase promoter as an example.
Won
Jaejoon
J
Biomedical Research Center, Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Daejeon, South Korea.
Kim
Tae Kook
TK
D000818:Animals; D002460:Cell Line; D047369:Chromatin Immunoprecipitation / Q000379:methods*; D003432:Cross-Linking Reagents / Q000494:pharmacology; D004247:DNA / Q000737:chemistry; D006657:Histones / Q000737:chemistry*; D006801:Humans; D009707:Nucleosomes / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D019098:Telomerase / Q000737:chemistry; D014157:Transcription Factors / Q000737:chemistry* / Q000378:metabolism; D014158:Transcription, Genetic
D003432:Cross-Linking Reagents; D006657:Histones; D009707:Nucleosomes; D014157:Transcription Factors; D004247:DNA; D019098:Telomerase