chip-Seq of mus musculus: es cell
In brief, approximately 1x107 cells were fixed with 1% formaldehyde at room temperature for 10 minutes and then quenched with 0.125 M glycine. The cross-linked cells were re-suspended in sonication buffer and sonicated using a Bioruptor. Sonication comprised three rounds with low mode pulsing settings (30s ON; 30s OFF), followed by ten rounds with high mode pulsing settings (30s ON; 30s OFF). A one-thirtieth fraction of the sonicated chromatin was preserved as input. The remaining chromatin was incubated with 1g of antibody conjugated to magnetic beads overnight at 4C. After the immunoprecipitation, the beads were washed consecutively with RIPA buffer, low salt buffer, high salt buffer, LiCl buffer, and 1X TE buffer. The DNA library was generated utilizing Tn5. Subsequently, DNA was extracted by reversing crosslinking at 65C overnight with proteinase K (20g/mL) and then sequenced using the Illumina HiSeq 2500 platform.