GSM3317769: HCT116 CDK11 ChIP G2 M phase; Homo sapiens; ChIP-Seq
Sample information curated by ChIP-Atlas
TFs and others
Cell type Class
Attributes by original data submitter
Abcam CDK11 (ab19393)
Sequenced DNA Library
ChIP was performed with Abcam CDK11 antibody (ab19393). Protein G Dynabeads (Thermo Fisher Scientific, 10009D) were pre-blocked with BSA for 4 h, washed 3 times with RIPA buffer (50 mM Tris-Cl, pH 8, 150 mM NaCl, 5 mM EDTA, 1% NP-40, 0.5% sodium deoxycholate, 0.1% SDS, supplemented with protease inhibitors, Sigma, P8340), followed by the incubation with specific antibody for at least 4 h at 4°C. Cells were crosslinked with 1% formaldehyde for 10 min, reaction was quenched with glycine (final concentration 125 mM) for 5 min. Cells were washed twice with ice-cold PBS, scraped, and pelleted. Each 20 µl packed cell pellet was lysed in 650 µl of RIPA buffer and sonicated 20 x 7s (amp 0.85) using 5/64 probe (QSonica Q55A). Clarified extracts (13,000g for 10 min) were precleared with protein G Dynabeads rotating for 2-4 h at 4°C and then incubated overnight with antibody pre-bound to protein G Dynabeads. For each ChIP-seq experiment (CDK11) we performed 3 technical replicates, dissolved each replicate in 17 µl of water and pooled them together to get at least 2.5 ng of immunoprecipitated DNA before library preparation (measured by Qubit). ChIP-seq libraries were generated using the KAPA Biosystems Hyper Prep Kit (KK8502) and NEBNext Multiplex Oligos for Illumina (Index Primers Set 1 and Set 2 (NEB E7335S, E7500S). Libraries were sequenced (50-bp single-end reads) using an Illumina HiSeq 2500 (VBCF Vienna).