GSM5129499: GATA1_186T_113G_revision; Homo sapiens; ChIP-Seq
2e7 HUDEP-2 cells were crosslinked with 1% formaldehyde for 10 minutes at room temperature. The reaction was quenched for 5 minutes at room temperature with a final concentration of 125 mM glycine. Fixed cells were lysed in 1mL Cell Lysis Buffer (10mM Tris-HCl pH 8, 10mM NaCl, 0.2% NP-40/Igepal). Nuclei were collected and lysed in 1mL Nuclear Lysis Buffer (50mM Tris-HCl pH 8, 10mM EDTA pH 8, 1% SDS). Chromatin was sonicated at 4 degrees C using a Branson 250 micro-tip sonicator with the power settings of 100% duty cycle, 10 second pulses for 2 minutes, with 90 seconds on ice between pulses. The sonicated chromatin was pre-cleared overnight at 4°C using protein A/G agarose beads. Pre-cleared chromatin was incubated with antibody. Following elution, DNA-protein complexes were treated with RNase for 30 minutes at 37°C followed by proteinase K treatment for 30 minutes at 45°C and overnight at 60°C with shaking. DNA was purified using a Qiagen MinElute kit. Libraries were prepared using standard reagents from New England BioLabs for Illumina sequencing