Characterization of in vivo DNA-binding events of plant transcription factors by chIP-seq: Experimental protocol and computational analysis

Abstract

Chromatin immunoprecipitation followed by next-generation sequencing (ChIP-seq) is a powerful technique for genome-wide identifi cation of in vivo binding sites of DNA-binding proteins. The technique had been used to study many DNA-binding proteins in a broad variety of species. The basis of the ChIP-seq technique is the ability to covalently cross-link DNA and proteins that are located in very close proximity. This allows the use of an antibody against the (tagged) protein of interest to specifi cally enrich DNAfragments bound by this protein. ChIP-seq can be performed using antibodies against the native protein or against tagged proteins. Using a specifi c antibody against a tag to immunoprecipitate tagged proteins eliminates the need for a specifi c antibody against the native protein and allows more experimental fl exibility. In this chapter we present a complete workfl ow for experimental procedure and bioinformatic analysis that allows wet-lab biologists to perform and analyze ChIP-seq experiments.