DNA Affinity Purification Sequencing (DAP-Seq) for Mapping Genome-Wide Transcription Factor Binding Sites in Plants

DNA affinity purification sequencing (DAP-seq) couples in vitro expression of transcription factor (TF) and binding to a genomic DNA library with next-generation sequencing, providing a robust method to identify genome-wide transcription factor binding sites (TFBS) on endogenous genomic DNA. Here we describe a detailed protocol of DAP-seq, including DNA library construction, cell-free protein expression, DNA affinity purification, and DNA sequencing. DAP-seq is an efficient and high-throughput approach to creating genome-wide binding site maps. Moreover, the genomic DNA library retains the chemical modifications on DNA such as cytosine methylation, allowing the effect of these modifications on TF binding to be analyzed. This method has been applied to the model plant Arabidopsis as well as cereal crops such as rice and maize.