Profiling of Transcription Factor Binding Events by Chromatin Immunoprecipitation Sequencing (ChIP‐seq)

Abstract In multi‐cellular organisms, gene expression is orchestrated by thousands of transcription factors (TF). Chromatin immunoprecipitation followed by sequencing (ChIP‐seq) is a robust tool to investigate gene expression because this technique profiles in vivo protein‐DNA interaction at a genome‐wide scale. Eight years after the first ChIP‐seq paper, there are limited reports of ChIP‐seq experiments in plants, especially for sequence‐specific DNA binding TFs. This lag greatly prevents our understanding of transcriptional regulation in an entire kingdom. In order to bridge the technical gap, we describe a ChIP‐seq procedure that we have successfully applied to dozens of sequence‐specific DNA binding TFs. The basic protocol includes procedures to isolate nuclei, sonicate chromatin, immunoprecipitate TF‐DNA complex, and recover ChIP‐enriched DNA fragments. The support protocol also describes practices to optimize library preparation by a gel‐free DNA size selection. Lastly, examples are given to optimize library amplification using real‐time PCR. © 2016 by John Wiley & Sons, Inc.