title: GSM5509519: D0_CTCF_K20R_rep2; Mus musculus; ChIP-Seq
library_construction_protocol: cells were crosslinked with 1% formaldehyde at room temperature for 10 min. Then the reaction was stopped by adding glycine (final concentration, 0.125 M). The cells were sonicated in SDS lysis buffer containing 1 x protease inhibitor cocktails and 1 mM PMSF to achieve a chromatin sized of 100-300 bp. The sonicated chromatin was incubated with indicated antibodies coupled with dynabeads protein A and G (1:1 mixed) at 4℃ overnight with rotation. Immune complexes were washed with the following buffers: low salt wash buffer (0.1% SDS, 1%Triton X-100, 2mMEDTA, 20mM Tris-HCl (pH 8.0), 150 mMNaCl), high salt wash buffer (0.1% SDS, 1% Triton X-100, 2mMEDTA, 20mM Tris-HCl (pH 8.0), 500 mM NaCl), LiCl wash buffer (0.25 MLiCl, 1% IGEPAL-CA630, 1% deoxycholic acid (sodium salt), 1 mM EDTA, 10 mM Tris-HCl (pH 8.0)) and TE buffer (10 mM Tris-HCl (pH 8.0), 1 mM EDTA). Antibody-bound chromatin was reverse-crosslinked, and the ChIPed DNA samples were purified for sequencing. ChIP DNA libraries were constructed following the Illumina ChIP-Seq library generation protocol.