ChIP-Seq Analysis    

Chromatin Immunoprecipitation (ChIP) is a method used to determine the location of DNA binding sites on the genome for a particular protein of interest. The ChIP assay offers great potential to improve knowledge about the regulation of gene expression. This technique is now used in a variety of life science disciplines including cellular differentiation, tumor suppressor gene silencing, and the effect of histone modifications on gene expression.

ChIP-sequencing (ChIP-Seq) combines chromatin immunoprecipitation (ChIP) and massively parallel sequencing to identify those DNA sequences bound by transcription factors in vivo.

A very common form of ChIP-Seq experiment is the Histone Modification Analysis. Histone compaction of DNA is a well-known epigenetic regulator of gene expression. Post-translational modification of histones, such as histone deacetylation, is another common form of epigenetic control, and is known to be associated with gene silencing. Therefore, evaluation of histone coverage and histone modification status are valuable tools in understanding genome-wide regulation of gene expression

How it works

Chromatin is first treated with formaldehyde to fix the chromatin-bound proteins to the DNA, followed by shearing into small fragment sizes (200bp – 1kb). This step is followed by immunoprecipitation using a specific ChIP-grade antibody. Crosslinking is reversed followed by proteinase K treatment or DNA purification using magnetic beads (IPure kit). The purified DNA is then submitted to ACGT, Inc. (See Important Considerations and Sample Submission Guidelines) and analyzed to identify the genomic regions where the specific DNA binding protein (e.g. histone) was located by generating a library using Illumina ChIP-Seq kit for analysis on the Genome Analyzer IIx.