GSM8156373: mESC_BPK25_HiC_rep1; Mus musculus; Hi-C
Genomic DNA were extracted. Four to five million cells were fixed for 5 min with 2 % formaldehyde in PBS and then the reaction was quenched for another 5 min in 0.125 M glycine. Nuclei were then extracted by incubating cells on ice for 30 min (inverting every 10 min) in 10 mM Tris pH 8.0, 10 mM NaCl, 0.2 % NP-40 (IGEPAL CA-630) and protease inhibitor cocktail (Roche).Hi-C libraries were prepared using an adaptor ligation method. Briefly, biotinylated junctions were captured on streptavidin M-280 dynabeads and digested with DpNII enzyme to generate blunt ends. The blunt ends were A-tailed with Klenow (3'→5' exo-) before ligation with adaptor sequences containing unique barcode. After adaptor ligation DNA was PCR amplified with Illumina primers for 25 cycles and library fragments of ~300-1000 bp (insert plus adaptor and PCR primer sequences) were isolated using AMPure beads (Beckman). The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Hiseq 4000 following the manufacturer's protocols.