Research assessing protein–DNA interactions at a genome-wide level can be used to understand chromatin dynamics and epigenetic modifications during normal development and disease. Isolation of protein–DNA interactions by chromatin immunoprecipitation (ChIP) followed by massively parallel DNA sequencing is an unbiased method for identifying and quantitating DNA sites that are associated to a chromatin-bound protein(s) of interest in any cell type or organism with known genomic sequence. Of particular interest are DNA regions involved in gene regulation, nucleosome positions, and epigenetic modifications to chromatin such as DNA methylation and histone modifications. For example, identifying transcription factor binding sites can be used to understand cell differentiation, environmental and drug responses, as well as alterations to these responses in disease states. Understanding the epigenetic modifications to chromatin is critical for determining the influence of chromatin states on transcription and the dynamic interplay between the epigenome and gene regulatory networks.
ChIP-Seq is an in vivo assay that uses antibodies to capture proteins that are bound to DNA directly or that interact within DNA-bound protein complexes. The ChIP-Seq workflow begins with formaldehyde treatment of live cells to crosslink proteins to DNA; this is followed by cell lysis and fractionation of the chromatin. An antibody specific to the protein of interest is used to immunoprecipitate the crosslinked protein–DNA complexes. The crosslinks are then reversed, and the resulting purified DNA is used in subsequent DNA fragment library construction for massively parallel DNA sequencing. Following sequencing, the sequence reads are analyzed using peak-calling tools that align reads to a reference sequence and identify putative DNA regions associated to proteins.
ChIP-Seq analysis provides fundamental insights, at a genome-wide level, into how chromatin is organized and regulated and how gene transcription is altered during many biological processes. Consider the following solutions to begin ChIP-Seq analysis in your lab:
The Ion Proton System combined with the MAGnify Chromatin Immunoprecipitation System delivers fast, reliable, and high-quality ChIP-Seq results at an affordable price.
For Research Use Only. Not for use in diagnostic procedures.