ATAC-seq Library Preparation Protocol

ATAC-seq libraries were generated as described [53 (link)]. Briefly stated, 50,000 cells were centrifuged, resuspended in 50 µL lysis buffer (10 mM Tris, 10 mM NaCl, 3 mM MgCl₂, and 0.1% IGEPAL CA-630), and centrifuged at 500× g for 10 min at 4 °C. The pellet was resuspended in transposase reaction mix (25 µL 2× TD buffer, 2.5 µL transposase (Nextera DNA sample preparation kit, Illumina), and 22.5 µL water, and incubated at 37 °C for 30 min. Tagmented DNA was purified using MinElute PCR Purification Kit (Qiagen, Hilden, Germany) per manufacturer’s instructions. DNA libraries were PCR-amplified using Nextera DNA sample preparation kit (Illumina, San Diego, CA, USA) using the following PCR conditions: 98 °C for 30 s, then thermocycling for 98 °C for 10 s, 63 °C for 30 s, and 72 °C for 1 min for 12 cycles, followed by 72 °C for 5 min. PCR products were size-selected for 200 to 800 base pair fragments using SPRI-Select Beads (Beckmann-Coulter, Brea, CA, USA). ATAC-seq reads were paired-end sequenced using an Illumina NextSeq500 (BSWRI Core Facility, Dallas, TX, USA).