NFATC1 ChIP-seq with antibody1
To induce nuclear localization of NFATC1 Jurkat cells were exposed to 2uM ionomycin and 100ng/ml PMA for twenty-four hours. Proteins were cross-linked to DNA by incubation of cells for 10 min in medium containing 1% formaldehyde at room temperature, after which the cross-linking was quenched, nuclei extracted and DNA fragmented by sonication. Precleared samples were incubated with 4 ug of specific antibody overnight at 4C and the antibodies collected by incubation with 30 ul of protein G- Sepharose beads for 2-3 h at  4C followed by centrifugation at 800 x g. Subsequently the beads were washed and precipitated chromatin complexes were eluted, and the cross-links reversed by incubating overnight at 65C. DNA was extracted with QiaQuick PCR purification kit