ChIP-Atlas: SRX527871

Antigen

library_strategy: ChIP-Seq
library_source: GENOMIC
library_selection: ChIP
library_construction_protocol: 10 million cells were crosslinked for 10 minutes in 1% formaldehyde followed by 5 minutes of quenching in 0.125 M glycine. Cells were then washed with cold PBS pH7.4, and collected in the presence of cold cell lysis buffer (5 mM PIPES pH 8.0, 85 mM KCl, 0.5% NP-40, and Roche Complete Protease Inhibitor Cocktail Cat #11836145001). Nuclei were pelleted by centrifugation at 2,000 rpm for 5 minutes at 4C, and the supertanant discarded. Sheared chromatin was then prepared by sonicating the nuclei in the presence of RIPA buffer (1 x PBS pH 7.4, 1% NP-40, 0.5% NaDOC, Roche Protease Inhibitor Cocktail), followed by centrifugation to remove cell debris. The chromatin was then immunoprecipitated with an anti-HA antibody (Covance MMS-101P). After immunoprecipitation, the formaldehyde crosslinks were reversed by heating overnight at 65C, and the DNA was collected using Qiagen PCR cleanup columns. DNA ends were prepared for Illumina HiSeq sequencing using the NEBNext Uktra DNA Library Prep Kit for Illumina (NEB Cat# E7370L). The adapted DNA was then size selected using SPRI beads for a size of 150-300bp, and PCR-amplified to make the final sequencing library. Samples were sequenced on an Illumina HiSeq 2000.