GSM3318539: NT-1-ATAC; Homo sapiens; ATAC-seq
ATAC-seq was performed on 50,000 nuclei. The samples were permeabilized in cold permeabilization buffer (0.2% IGEPAL-CA630 (I8896, Sigma), 1 mM DTT (D9779, Sigma), Protease inhibitor (05056489001, Roche), 5% BSA (A7906, Sigma) in PBS (10010-23, Thermo Fisher Scientific)) for 10 minutes on the rotator in the cold room and centrifuged for 5 min at 500 xg at 4°C. The pellet was resuspended in cold tagmentation buffer (33 mM Tris-acetate (pH = 7.8) (BP-152, Thermo Fisher Scientific), 66 mM K-acetate (P5708, Sigma), 11 mM Mg- acetate (M2545, Sigma), 16% DMF (DX1730, EMD Millipore) in Molecular biology water (46000-CM, Corning)) and incubated with Tagmentation enzyme (FC-121-1030; Illumina) at 37 °C for 30 min with shaking 500 rpm. The tagementated DNA was purified using MinElute PCR purification kit (28004, Qiagen). The libraries were amplified using NEBNext High-Fidelity 2X PCR Master Mix (M0541, NEB) with primer extension at 72°C for 5 minutes, denaturation at 98°C for 30s, followed by 8 cycles of denaturation at 98°C for 10 s, annealing at 63°C for 30 seconds and extension at 72°C for 60 seconds. After the purification of amplified libraries using MinElute PCR purification kit (28004, Qiagen), the double size selection was performed using SPRIselect bead (B23317, Beckman Coulter) with 0.55X beads and 1.5X to sample volume.