GSM8048444: ATAC_SMANTIS_KO_3; Homo sapiens; ATAC-seq
50,000 THP-1 were used for the ATAC library preparation performed with the Illumina Tagment DNA Enzyme and Buffer Kit (20034198, Illumina). Cells were washed in 50 µl cold PBS and centrifuged at 1200 rpm for 4 min. The cell pellet was resuspened in 50 µl of cold lysis buffer (48.5 µl of resuspension buffer (10 mM Tris-HCl pH 7.5 (15567-027, Thermo Fisher Scientific), 10 mM NaCl, 3 mM MgCl2), 0.5 µl 10% NP-40, 0.5 µl 10% Tween-20 (P1379, Sigma-Aldrich), and 0.25 µl 2% Digitonin (G9441, Promega)) and incubated for 3 min on ice. Wash buffer (990 µl of resuspension buffer, 10 µl 10% Tween-20) was added to the cell lysate and centrifuged at 1000 xg at 4°C for 5 min. The cell pellet was resuspened in transposition reaction mix (25 µl TD Buffer, 16.5 µl PBS, 0.5 µl 10% Tween-20, 0.25 µl 2% Digtonin, 2.5 µl Tn5, 5 µl H2O) and incubated at 1000 rpm at 37°C for 30 min. DNA was purified by using MinElute PCR Purification Kit (Qiagen), eluated in 20 µl elution buffer, and stored at -20 °C until sequencing. The amplification of the library together with indexing primers was performed as described by Buenrostro et al. 2013. Libraries were mixed in equimolar ratios and sequenced on NextSeq2000 platform using a P3 flowcell ATAC-seq was performed based on the protocol from Buenrostro et al. 2013 and modified by Ackermann et al. 2016.