title: GSM2808031: K562_A1_gH2Ax; Homo sapiens; ChIP-Seq
library_construction_protocol: Sequencing libraries were constructed from DNA samples with the Illumina TruSeq V3 library construction protocol. Cells were treated as indicated, crosslinked with 1% formaldehyde in PBS for 10 min, followed by quenching with 125 mM glycine. Cells were then resuspended in lysis buffer (5 mM PIPES pH 8.0, 85 mM KCl, 0.5% nonidet P-40) to isolate nuclei. Nuclei were resuspended in micrococcal nuclease (MNase) digestion buffer (10 mM Tris pH 7.4, 15 mM NaCl, 60 mM KCl, 0.15 mM spermine, 0.5 mM spermidine) and 1.2 U/µl MNase was added for 30-45 min at 37 °C. The reaction was stopped by adding 50 mM EDTA and nuclear pellets were resuspended in 10mM Tris-HCl (pH8.0), 100 mM NaCl, 1mM EDTA, 0.5mM EGTA, 0.1% Na-deoxycholate, 0.5% N-lauroylsarcosine. Lysates were sonicated briefly to disrupt nuclear membranes using an ultra sonicator water bath (Bioruptor, Diagenode). Diluted lysates were incubated o/n at 4 °C with the indicated antibodies after addition of 1% Triton X-100. IPs were performed using 30 µl Protein A/G magnetic beads (Pierce). Eluted DNA was purified with QIAquick PCR purification (Qiagen) according to the manufacturer instructions. Purified ChIP DNA was analyzed by qPCR using a LightCycler 480 II (Roche). Sequencing libraries were constructed from DNA samples (ChIP and control samples) with the Illumina TruSeq V3 library construction protocol (Illumina, San Diego, USA).
attributes.tag: GEO Accession
attributes.value: GSM2808031