GSM5628881: ChIP_mouse_WT_UNSTIM_1_H3K27ac_IP; Mus musculus; ChIP-Seq
DNA and proteins from sorted NK cells were cross-linked for 8 minutes at room temperature using 1% formaldehyde. Cross-linked cells incubated in cell lysis buffer (25 mM HEPES, 1.5 mM MgCl2, 10mM KCl, 0.1% NP-40, 1mM DTT, 1x proteinase inhibitor) and nuclei were isolated by centrifugation at 5400 rpm for 5.5 minutes. ChIP was performed using 1.5 μg of rabbit polyclonal anti-trimethyl-Histone H3 (Lys4) antibody (H3K4me3, Millipore, 07473) or 8 µg anti-Stat5 (#AF2168, R&D Systems) and Dynabeads Protein G (Invitrogen). Immunoprecipitated DNA was quantified by PicoGreen and the size was evaluated by Agilent BioAnalyzer. Whenever possible, fragments between 100 and 600 bp were size selected using aMPure XP beads (Beckman Coulter catalog # A63882) and Illumina libraries were prepared using the KAPA HTP Library Preparation Kit (Kapa Biosystems KK8234) according to the manufacturer's instructions with up to 10ng input DNA and 8-11 cycles of PCR. Barcoded libraries were run on a HiSeq 2500 in Rapid or High Output mode or a HiSeq 4000 in a 50bp/50bp paired end run, using the HiSeq Rapid SBS Kit v2, HiSeq SBS Kit v4, or HiSeq 3000/4000 SBS Kit, respectively (Illumina). An average of 46 million paired reads were generated per sample.