GSM5059701: MV411_MEF2D_ChIP; Homo sapiens; ChIP-Seq
In brief, 20 million MOLM-13 or MV4-11 cells were cross-linked with formaldehyde (1%) for 10 minutes followed by quenching with 0.125 M glycine for 5 minutes at room temperature. After washing with cold PBS, cells were suspended in lysis buffer (50 mM HEPES pH 7.5, 150 mM NaCl, 1 mM EDTA, 0.5 mM EGTA, 1% TritonX-100, 0.1% SDS, 0.1% N-lauroylsarcosine, and protease inhibitor) and subjected to sonication to shear the chromatin DNA into sizes ranging from 200 to 500 bp. The fragmented chromatin was pre-cleared by centrifugation at 14,000 rpm for 15 min at 4°C. The supernatant was incubated with primary antibody and Protein A or G Dynabeads magnetic beads (Invitrogen) at 4°C overnight. After four times wash with high-salt buffer (20 mM Tris-HCl pH 8.0, 2 mM EDTA, 0.1% SDS, 500 mM NaCl, and 1% TritonX-100) and twice wash with TE buffer (10 mM Tris-HCl pH 8.0, 1 mM EDTA), chromatin was eluted from beads in elution buffer (50 mM Tris-HCl pH 8.0, 10 mM EDTA, 1% SDS). After reverse crosslinking, ChIP DNA was purifiied using DNA purification columns (Zymo Research) and subjected to downstream quantitative real-time PCR analysis (ChIP-qPCR) or library preparation for deep sequencing (ChIP-seq). ChIP-seq library was prepared using NEBNext Library Prep Kit following the instructions and subjected to paired-end, 150-cycle sequencing on the Illumina HiSeq 4000 platform.