title: GSM2481675: KK1 BATF3; Homo sapiens; ChIP-Seq
library_construction_protocol: BATF3 Antibody Extraction Other: 20 million exponentially growing cells were collected per sample and cross-linked with 1% formaldehyde for 10 min at room temperature. The cross-linked cells were resuspended in ice-cold RIPA buffer (10mM Tris-HCl pH8, 140 mM NaCl, 1mM EDTA pH 8, 0.5 mM EGTA, 1% Triton X-100, 0.1% SDS and 0.1% sodium deoxycholate) to a final concentration of 4 million cells/ml. DNA was sheared with a Misonix XL sonicator. For each immune precipitation reaction, the chromatins were incubated overnight with 5 ug of BATF3 antibody (R&D, AF7437).The following day, chromatin/antibody complexes were incubated with Protein G Dynabeads (50 ul; Thermo Fisher Scientific) for 4 h at 4 C, washed 3 times with RIPA Buffer, once with LiCl Buffer (10 mM Tris-HCl pH8, 250 mM LiCl, 0.5% NP40, 0.5% Sodium Deoxycholate, 1 mM EDTA), once with TE pH 8.0 and finally resuspended in 100 ul TE pH8 containing RNase A (0.2 ug/ul). Reverse crosslinking was performed overnight at 65 C, followed by treatment with 20 ug Proteinase K (Invitrogen) for 2 h at 50 C. Final DNA purification was performed with QIAquick PCR Purification columns (QIAGEN). ChIP DNA was used to generate ChIP-seq libraries with the NEXTflex Illumina ChIP-seq Library Prep Kit (Bio Scientific) according to manufacturer's instructions.
identifiers.namespace: GEO
identifiers.id: GSM2481675