Chromatin accessibility assay

Five thousand cells were isolated from each sample and lysed in 5 to 10 μl of ice-cold ATAC lysis buffer (10 mM Tris-HCl, 10 mM NaCl, 3 mM MgCl₂, and 0.1% IGEPAL CA-630 pH 7.5) for 10 min. The nuclear extraction step was skipped and Illumina’s adaptors were ligated using the transposase reaction mix of Nextera DNA Sample Prep Kit (Illumina), and the samples were incubated for 30 min at 37 °C. Indices were incorporated using Nextera Index Kit (Illumina) in the amplification step. The libraries were purified using Ampure XP beads (Beckman Coulter) to remove remaining adapters. The libraries were paired-end sequenced on Illumina HiSeq 2500 for 64 cycles. Sequenced reads were handled as described in ChIP-seq data analysis above, except that the reads mapping to the mitochondrial DNA (chrM) were excluded for further analysis. ATAC-seq peak sites were defined as sites detected in two independent experiments.