Chromatin immunoprecipitation of skeletal muscle tissue

Abstract

Chromatin immunoprecipitation (ChIP) is an invaluable method for studying interactions between histone proteins and genomic DNA regions and transcriptional regulation using antibodies to enrich genomic regions associated with these epitopes. Either to monitor the presence of histones with post-translational modifications at specific genomic locations or to measure transcription factor interactions with a candidate target gene, protein–DNA complexes are most commonly crosslinked using formaldehyde, which stabilizes these transient interactions. Chromatin is then fragmented to allow separation of genomic fragments bound by the histone or transcription factor of interest away from those that are unbound. Following immunoprecipitation, formaldehyde crosslinks are reversed and enriched DNA fragments are purified. While some investigators have successfully performed ChIP experiments from crosslinked skeletal muscle in cell culture, the process is relatively inefficient compared to whole tissue. This chapter provides protocols specifically designed for the crosslinking and immunoprecipitation of human skeletal muscle biopsy samples in preparation for chromatin immunoprecipitation-sequencing (ChIP-seq).