GSM1402793: Cit_10min; Homo sapiens; ChIP-Seq
To achieve high temporal resolution, ChIP crosslinking concentration of formaldehyde was 2% and crosslinking was quenched after 1 minute with 250mM glycine. The H3R26Cit antibody we used for IP was AbCam: ab19847. The H3K27ac antibody we used for IP was Millipore: 07-360. The ends of the ChIP DNA were blunted and phosphorylated in 1X END Repair Buffer (Epicentre Biotechnologies, #ER0720), 250nM dNTPs, 1mM ATP, and 1X END-IT enzyme mix (Epicentre Biotechnologies, #ER0720) for 45 minutes. An A overhang was achieved in 1xNEB2 buffer, 200nM dATP (Invitrogen), 3units of Klenow (3¢-5¢ exo-) 5U/ml (from New England BioLabs), in 50ul volume for 30 minutes. 10nM of adapter was ligated to the DNA in 1X ligation buffer, 5ml (3000units) of UltraPure Ligase from enzymatics (Enzymatics T4 DNA Ligase #L603-HC-L) in a total volume of 50ul. Thirteen cycles of PCR with Phusion polymerase were performed to acquire the final library.
GEO Accession
GSM1402793