NIH 3T3 ATAC-seq Protocol for Chromatin Profiling

For NIH 3T3 ATAC-seq^{75 (link)}, 100,000 cells were used per experiment and for liver samples 3 mg of pulverized tissue was used as starting material. Following centrifugation at 500 g for 5 min at 4 °C, cells were washed with cold PBS and then resuspended in cold lysis buffer (10 mM Tris-HCl, pH 7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% NP-40). Cells were lysed on ice for 15 min and then spun down at 500 g for 10 min at 4 °C. After removing the supernatant, the nuclei were resuspended in transposition reaction mix (25 µl TD 2x reaction buffer, 2.5 µl Nextera Tn5 Transposase, 22.5 µl nuclease free water). The reaction was incubated for 30 min at 37 °C. DNA was purified using a PCR purification kit (ThermoFisher) and eluted in elution buffer (10 mM Tris buffer, pH 8). DNA was amplified for 10 cycles using a thermocycler and barcoded primers^{74 (link)}. DNA purification was achieved using sparQ PureMag Beads (Quantabio). Libraries were sequenced at the University of British Columbia (now SBME Sequencing Facility).