title: GSM5896815: ATAC-seq_G7FSen_2; Mus musculus; ATAC-seq
library_construction_protocol: Sorted cells were directly processed for ATAC-seq library preparation without in vitro culture.  Sorted cells were centrifuged at 500 × g for 5min at 4°C and lysed using cold ATAC-RSB (10mM Tris-HCl, pH7.4,10mM NaCl, 3mM MgCl2 supplemented in 0.1% NP40, 0.1% Tween20 and 0.1mg/ml Digitonin) on ice for 3min. Next, the lysed cells were added in 50µl of transposition mix (25 µl of 2 × TD buffer (20mM Tris-HCl, pH7.6, 10mM MgCl2, 20% Dimethyl Formamide), 1.5 µl transposase (in house, (Picelli et al., 2014)), 17.5µl of PBS, 0.5µl of 10% Digitonin, 0.5µl of 10% Tween20 and 5µl of nuclease-free water). The transposition reaction was carried out for 30 min at 37°C and placed at 10°C. The transposition sample was purified using a Qiagen MinElute kit and eluted using 21µl of EB buffer. The transposed fragments were amplified using 50ul of PCR mix (20µl of DNA, 2.5µl of custom Nextera PCR primers 1 and 2, and 25µl of KAPA HiFi HS Ready Mix for a total of 15 cycles. The PCR amplification conditions were 72°C for 5 min and 95°C for 30 sec, followed by 15 cycles at 95 °C for 10 s, 63 °C for 30 s, and 72 °C for 60 s, with a final extension at 72 °C for 5 min. Following PCR amplification, the libraries were purified, and the size was selected from 150 to 800bp using AMPure XP beads.