title: GSM8027061: EedKO_H3K9me3; Mus musculus; ChIP-Seq
library_construction_protocol: ~1x106 cells were fixed in 1% formaldehyde for 25 min at room temperature immediately after isolation. The reaction was stopped by addition of Glycine to a final concentration of 0.13 M.  Cells were lysed using a buffer containing 30 mM Tris-HCl (pH 8), 1% SDS, 10 mM EDTA, and protease inhibitors (Roche), and the chromatin was sonicated for 50 min using a Covaris sonicator (5-min on/off cycles at 4 °C). Debris were removed using centrifugation and the chromatin was incubated overnight at 4°C with antibody against H3K9me3 (Abcam, Ab8898). Antibody-bound chromatin was captured using magnetic beads (Themo Fisher, 10001D) and washed using low-salt (20 mM Tris-HCl pH 8.1, 150 mM NaCl, 2 mM EDTA, 0.1% SDS, 1% TritonX-100), high-salt (20 mM Tris-HCl pH 8.1, 500 mM NaCl, 2 mM EDTA, 0.1% SDS, 1% TritonX-100), and lithium chloride (10 mM Tris pH 8.1, 0.25 M LiCl, 1 mM EDTA, 1% NP-40, 1% deoxycholate) buffers. Chromatin was treated with 1% SDS and 0.1 M NaHCO3 for 6 h at 65°C to reverse the cross-links and DNA was purified using columns Illumina sequencing libraries were generated by end repair, 3' A-addition, and Illumina sequencing adaptor ligation.