title: GSM4875721: NC3_ctrl_ATAC-Seq_rep2; Homo sapiens; ATAC-seq
library_construction_protocol: ATAC-seq was performed using the OMNI-ATAC protocol Briefly, cells were dissociated using Accutase, and 50,000 cells were subjected to the tagmentation reaction. Cells were first washed in resuspension buffer (10 mM Tris-HCl pH 8.0, 10 mM NaCl, and 3 mM MgCl2 in water), following which nuclei were isolated in 1 ml lysis buffer (10 mM Tris-HCl pH 8.0, 10 mM NaCl, 3 mM MgCl2, 0.1% NP-40, 0.1% Tween-20, and 0.01% Digitonin in water) on ice for three minutes. Nuclei were rinsed once in wash buffer (10 mM Tris-HCl pH 8.0, 10 mM NaCl, 3 mM MgCl2, and 0.1% Tween-20) and tagmentation was carried out using 2.5ul Tn5 transposase (Illumina 15027865) for 30 mins. Following tagmentation, DNA was purified using the Zymo DNA Clean and Concentrator kit. Libraries were prepared by PCR using Q5 High Fidelity DNA polymerase (NEB) polymerase and using primers carrying Illumina Nextera i7 barcodes. First, gap filling was performed at 72°C for 5mins followed by five cycles of 98°C, 20secs, 63°C, 30secs, and 720C 1min. After initial amplification, tubes were held on ice, while quantitative PCR was run on 1l of the pre-amplified library to determine additional number of cycles needed. Libraries were sequenced on HiSeq2500 using PE50.