GSM3958116: HEK293-mock_antiHA-Ab; Homo sapiens; ChIP-Seq
HEK293 cells expressing ZFAT-HA were crosslinked with 1% formaldehyde for 10 min at room temperature. Crosslinking reaction was quenched with 125 mM glycine for 5 min. The cells were rinsed with PBS twice and incubated in 600 µl of MNase treated buffer (50 mM Tris-HCl, pH 8.0, 150 mM NaCl, 0.1% NP-40, 5 mM CaCl2) containing 375 unit MNase (Roche) for 0.5 h at 37°C. The MNase reaction was stopped by adding 600 µl of lysis buffer  (100 mM Tris-HCl, pH 8.0, 20 mM EDTA, 2% SDS, protease inhibitor cocktail (Roche)), and the lysate was sonicated using a Bioruptor (Cosmo Bio, Tokyo, Japan) for two cycles of one minute with 30 seconds on/off. The sonicated lysate was subjected to immunoprecipitation with a monoclonal antibody against HA-tag or control IgG. The immunoprecipitated DNA was purified using MinElute Reaction Cleanup kit (Qiagen) according to the manufacturer's instructions. ChIP-seq libraries were prepared using NEBNext ChIP-seq Library Prep Master Mix Set and Multiplex Oligos for Illumina (New England Biolabs) according to manufacturer's instructions. ChIP or input DNA (0.1ng) was subjected to end repair, dA-tailing, and adaptor-ligation, and amplified by 13 cycles of PCR.
GEO Accession
GSM3958116