title: GSM5366626: KDM4A-OE.ATAC.G1.rep1; Homo sapiens; ATAC-seq
library_construction_protocol: Sorted cells were pelleted and resuspended in 250 ml cold RBS buffer (10 mM Tris-HCl pH 7.4 – 10 mM NaCl – 3 mM MgCl2) before being pelleted by centrifugation. Pellet was resuspended in 250 ml cold RBS buffer – 0.1% NP-40, incubated 5 minutes on ice and pelleted by centrifugation. Pellet was reuspended in 50 ml Tagmentation mix (22.5 ml H2O – 25 ml 2X Tagment DNA buffer – 2.5 ml Tagment DNA enzyme, Nextera Illumina FC-121-1030) and incubated 30 minutes at 37 degrees before being placed ion ice. Protocol was adapted from (Buenrostro et al Nat Methods 2013). Purified DNA was amplified by PCR by adding 2.5 ml 25 mM Primer Ad2 and 37.5 ml PCR mix (10 ml H2O – 2.5 ml 25 mM Primer Ad1 – 25 ml 2X NEBnext master mix, NEB E6040S) and incubated as follows: 72 degrees 5 minutes, 98 degrees 30 seconds, [98 degrees 10 seconds, 63 degrees 30 seconds, 72 degrees 1 minute] x 12 cycles, 4 degrees. Amplified DNA was purified using a Promega Gel Purification Kit following supplier's instructions and eluted in 25 ml H2O. Library length distribution were assessed by TapeStation (Agilent) and quantified with Qubit (ThermoFisher). Libraries were paired-end sequenced (43 cycles each way) using a NextSeq500 (Illumina).