GSM5255998: human PB NK cells RUNX2 Input; Homo sapiens; ChIP-Seq
Sorted human peripheral blood NK cells were fixed in 1% formaldehyde and quenched with 0.125M glycine. DNA was sheared by sonicating the lysates with a microtip sonicator. Genomic DNA (Input) was prepared by treating aliquots of chromatin with RNase, proteinase K and heat for de-crosslinking, followed by ethanol precipitation. For each ChIP reaction, 35.5 ug of chromatin was precleared with protein A agarose beads (Invitrogen). ChIP reaction was set up using precleared chromatin and antibody and incubated overnight at 4°C. Protein A agarose beads were added and incubation at 4°C for an additional 3 h. Immune complexes were washed, eluted from the beads with SDS buffer, and subjected to RNase and proteinase K treatment. Crosslinks were reversed by incubation overnight at 65°C, and ChIP DNA was purified by phenol-chloroform extraction and ethanol precipitation. Illumina sequencing libraries were prepared from the ChIP and Input DNAs by the standard consecutive enzymatic steps of end-polishing, dA-addition and adaptor ligation. After a final PCR amplification step, the resulting DNA library was quantified.