KAS-Seq Maps Transcriptional Dynamics and Active Enhancers by Examining Single-Stranded DNA Landscapes

Genome-wide analysis of transcription activity and its regulation is critical for understanding cell functions. We have recently developed kethoxal-assisted single-stranded DNA sequencing (KAS-seq) and shown that ssDNA in coding regions can serve as a precise readout of transcriptional activity. Compared with ChIP-seq and nascent RNA sequencing, KAS-seq has a simpler procedure that works with low-input materials. KAS-seq reveals transcriptional dynamics with high temporal resolution and maps enhancer activity in the same assay. This chapter describes a detailed protocol for labeling, enriching, and sequencing ssDNA (KAS-seq) to assay transcription and enhancer activities in cultured mammalian cells.