Chromatin immunoprecipitation (ChIP) reveals the dynamics of histone modifications and transcription co-factors, bound to specific genes, or their binding in genome-wide profiles using microarray or sequencing (1). ChIP data always represents a population of nucleosomes, from cultured cells or tissues. The method is dependent on the global abundance of any histone modification targeted and the affinity of the antibodies used. Samples can be prepared to compensate for these limitations (2). Cross-linking ChIP or X-ChIP uses formaldehyde fixation of chromatin complexes bound to DNA. Nuclei are then lysed and the DNA is sheared and co-immunoprecipitated with antibodies to histones or histone modifications. Next, the crosslinking is reversed and DNA is purified for downstream analysis. Native ChIP (NChIP) is an otherwise similar method, but uses chromatin prepared by micrococcal nuclease digestion of sample cell nuclei. NChIP has higher resolution data. However, it is more suitable for histones and less suitable for fragile DNA-binding by other chromatin complex protein targets. Various ChIP methods are applied by researchers in developing a “Histone Code” which is hypothesized as the combinations of histone modifications that would predict changes in gene expression (3). Another variation of ChIP method in combination with exonuclease I (ChIP-exo) is capable of mapping the locations at which a protein of interest (transcription factor) binds to the genome (4).
Protocols for ChIP
ChIP-Seq Library Construction Workflow
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If all cells are created from the same genetic material, why are there so many different cell types? Listen to Sriharsa Pradhan, Senior Scientist, RNA Biology at NEB, as he describes how DNA is methylated and how this affects the path of reading the DNA code the same way an obstruction would derail a train off its tracks.