GSM3781511: MOLM13 cells, R248Q, DMSO; Homo sapiens; ChIP-Seq
20x10e6 cells were crosslinked using 1% methanol-free formaldehyde (ThermoFisher, #28906) in PBS for 7 min at room temperature. Remaining formaldehyde was quenched with 100 mM Tris-HCl pH8.0 and 25 mM Glycine. Cytoplasm was stripped using 50 mM Tris-HCl pH 8.0, 100 mM NaCl, 5 mM EDTA, 1% SDS for 10 min at room temperature followed by precipitation of nuclei by centrifugation at 10,000g. The nuclei were then resuspended in 66 mM Tris-HCl pH 8.0, 100 mM NaCl, 5 mM EDTA, 1.7% Triton X-100, 0.5% SDS and sheared using an E220 sonicator (Covaris). Chromatin fragmentation was quality-controlled using ScreenTape D5000 (Agilent). Immunoprecipitation with anti-p53 antibodies (DO-1, abcam, #1101) and Protein G Dynabeads™ (ThermoFisher, #10003D) was performed overnight at 4 degree Celsius. Beads were washed and immunoprecipitated DNA was reverse-сrosslinked in 100 mM NaHCO3, 100 mM NaCl, 1% SDS, and quantified by ScreenTape HS D1000 (Agilent) and Qubit (ThermoFisher). Illumina-compatible sequencing libraries were prepared using SMARTer® ThruPLEX® DNA-Seq Kit (Takara, #R400675) and sequenced using a NextSeq Sequencing System (Illumina) to obtain paired-end reads.