GSM5393762: E14-NT; Mus musculus; ChIP-Seq
Lentivirus transduced ESCs were fixed using 1% formaldehyde for 10 min, and 0.125 M glycine was added to stop the fixation. Cells were then harvested, and DNA was fragmented to 300–500 base pairs (bp) by sonication with Scientz08-III sonicator (SCIENTZ). Immunoprecipitation was performed with antibodies (Pol II, Santa Cruz, sc-47701) conjugated to Dynabeads protein A/G beads (Pierce). The ChIPped DNA was eluted, reverse-cross-linked, extracted by AMPure XP beads (Beckman). Libraries were prepared according to the instructions accompanying the NEBNext Ultra II DNA Library Prep Kit for Illumina (NEB). Briefly, DNA was end-repaired and treated with Klenow fragment to yield a protruding 3- 'A' tail for ligation of Illumina's adapters.Then, DNA was amplified with Illumina primers for 13 cycles using PCR. PCR products were purified with AMPure XP beads (Beckman). The libraries were sequenced on the Illumina NovaSeq 6000 following the manufacturer's protocols.