hFetal_Input1_InputChIPH3K27ac_R01L01S01
For H3K27ac, 5-15 x 10^6 formaldehyde crosslinked iPSC-RPE cells were lysed and sonicated in 110 ul of SDS Lysis Buffer (0.5% SDS, 50 mM Tris-HCl pH 8.0, 20 mM EDTA, 1x cOmplete Protease Inhibitor Cocktail (Sigma)) using Covaris E220 Focused-ultrasonicators (Covaris) for 14 cycles, 1 min per cycle, duty cycle 5. Next, samples were reverse crosslinked by incubation for 15 min at 65C with rotation and subsequent incubation with 5 ul RNAse (Sigma) for 1h at 37C and Proteinase K Solution (20 mg per mL, Thermo Fisher Scientific) for 1h at 55C. After reverse crosslinking, samples were purified with 2X Agencourt AMPure XP DNA beads (Beckman Coulter), eluted in 30 ul of H2O, and quantified using Qubit (Thermo Scientific). Libraries were generated using KAPA Hyper Prep Kit (KAPA Biosystems) and KAPA Real-Time Library Amplification Kit (KAPA Biosystems) following the manufacturer's manual. Libraries were barcoded using TruSeq RNA Indexes (Illumina) and sequenced on an Illumina HiSeq 4000 to an average of 40 M 150 bp paired-end reads per sample.