GSM8603258: Runx3-ChIP, Runx3 +/+, rep.1; Mus musculus; ChIP-Seq
Freshly isolated thymocytes were cross-linked with 1% formaldehyde. Nuclei were isolated from the cells using lysis buffer and chromatin DNA was fragmented by sonication. The lysate was incubated with rabbit anti-mouse Runx3 (Cell Signaling Technology, D6E2), anti-H3k4me3 (Cell Signaling Technology, C42D8), or anti-H3K27me3 (Cell Signaling Technology, C36B11) antibodies pre-bound to Dynabeads® M-280 Sheep anti-Rabbit IgG overnight at 4 degree C. Chromatin DNA was eluted from Dynabeads and reverse-crosslinked. Samples were treated with RNase A and Proteinase K, and DNA was purified by phenol/Chroroform/Isoamylalcohol treatment and echanol precipitation. Purified DNA was subjected to the re-sonication with a Covaris S220 to produce DNA fragments with an average size of 200 bp, and was used for library preparation with NEBNext ChIP-seq Library Prep Master Mix Set for Illumina and NEBNext Multiplex Oligos for Illumina (New England BioLabs). Sequencing was performed by genomics facility at RIKEN IMS with Illumina HiSeq 2500.