iPSC H3K27ac ChIP-Seq: Sample cfb4725b-02ae-40b0-b4c9-06edd4393edc
We performed H3K27AC ChIP-Seq in 52 iPSC samples from 46 lines from 36 individuals from iPSCORE. Pellets of formaldehyde -crosslinked iPSCs were lysed and sonicated in 110 µl of SDS Lysis Buffer (0.5% SDS, 50mM Tris-HCl pH 8.0, 20mM EDTA, 1x cOmplete™ Protease Inhibitor Cocktail (Sigma)) using Diagenode Bioruptor UCD-200 (Diagenode) or Covaris E220 Focused-ultrasonicators (Covaris). For each sample, 1 µg of H3K27ac antibody (Abcam ab4729) was coupled for 2-4h to 11µl of Protein G Dynabeads (Thermo Scientific) and used for overnight chromatin immunoprecipitation in IP buffer (1% Triton X-100, 0.1% DOC, 1x TE buffer, 1x cOmplete™ Protease Inhibitor Cocktail). 40-45µg of chromatin (66ug  1 sample) was used for immunoprecipitation. Beads with immunoprecipitated chromatin were washed five times with 150 µl of RIPA buffer (50mM HEPES pH 8.0, 1% NP-40, 0.7% DOC, 500mM LiCl, 1mM EDTA, 1x cOmplete™ Protease Inhibitor Cocktail) and once with 1X TE buffer (10mM Tris-HCl pH 8.0, 1mM EDTA). Next samples were eluted in 150 µl of ChIP Elution Buffer (1% SDS, 10mM Tris-HCl pH 8.0, 1mM EDTA) and reverse crosslinked by incubation for overnight at 65°C and subsequent incubation with 5 µl RNAse (Sigma) for 1h at 37°C and Proteinase K Solution (20 mg/mL, Thermo Fisher Scientific) for 1h at 55°C. After reverse crosslinking, samples were purified with MIniElute PCR purification kit (Qiagen) or with DNA Clean & Concentrator kit (Zymo), eluted in 25µl of EB buffer (Qiagen)and Qubit (Thermo Scientific) quantified. Libraries were generated using KAPA Hyper Prep Kit (KAPA Biosystems) and KAPA Real Time Library Amplification Kit (KAPA Biosystems) at Institute for Genomic Medicine at University of California, San Diego. Libraries were barcoded using TruSeq RNA Indexes (Illumina). Libraries were sequenced on an Illumina HiSeq 4000 100bp Paired-End reads. Peaks were called using MACS2 --broadpeak on all samples simultaneously.
alignment_software
BWA-MEM