GSM8606434: LNCaP_INPUT; Homo sapiens; ChIP-Seq
The cells were first fixed at room temperature with 2mM DSG (Thermo Scientific, Cat.No. 20593) for 45 min prior to 1% formaldehyde fixation for 10 min. The cells sequentially lysed with LB1 (50 mM HEPES-KOH (pH 7.6), 140 mM NaCl, 1 mM EDTA, 10% (vol/vol) glycerol, 0.5% (vol/vol) NP-40/Igepal CA-630 and 0.25% (vol/vol) Triton X-100), LB2 (10 mM Tris-HCl (pH 8.0), 200 mM NaCl, 1 mM EDTA and 0.5 mM EGTA) to enrich for nucleus. The nucleus was further lysed in LB3 and sonicated for 10 mins (30 sec 'On' and 30 sec 'Off' cycles) using Bioruptor (Diagenode). The IP was performed overnight at 4°C with 4 µg of BRG1 (Abcam, ab110641) or SMARCC2 (Bethyl Lab A301-039A) conjugated protein G magnetic beads (Thermo Scientific). The DNA was then purified using the Monarch DNA cleanup kit (NEB) after overnight de-crosslinking at 65°C. The library for high throughput sequencing was prepared using the Ultra II DNA library kit (NEB) according to the manufacturer's instructions.