GSM9684138: H3K27ac_CD2_cntrl_rep2; Homo sapiens; ChIP-Seq
Cells were lysed as described in Ortabozkoyun et al., 2022 using buffers in the following order: LB1 (50 mM HEPES, pH 7.5 at 4 C, 140 mM NaCl, 1 mM EDTA, 10% Glycerol, 0.5% NP40, 0.25% Triton X) for 10 min at 4 C, LB2 (10 mM Tris, pH 8 at 4 C, 200 mM NaCl, 1 mM EDTA, 0.5 mM EGTA) for 10 min at RT, and LB3 (10 mM Tris, pH 7.5 at 4 C, 1 mM EDTA, 0.5 mM EGTA, and 0.5% N-Lauroylsarcosine sodium salt). Chromatin was sonicated to an average size of ~250 bp using a Covaris sonicator. ChIP was performed by using antibodies listed. Chromatin from Drosophila (1:100 ratio to CD2 cells chromatin), and Drosophila-specific H2Av antibody were used as spike-in control in each sample. ChIP-seq Libraries were prepared according to manufacturer's instructions (Illumina) as described in Narendra et al., 2015. Immunoprecipitated DNA was first end-repaired using End-It Repair Kit (Epicenter), tailed with an A using Klenow exo minus (NEB M0212), ligated to custom adapters, and fragments of 300±100 bp were size-selected. These fragments were amplified through ligation-mediated PCR amplification (LM-PCR) by using Q5 polymerase (NEB M0530).