GSM4212721: pooled HiC; Homo sapiens; Hi-C
The cells were washed with PBS and then harvested in Hi-C lysis buffer (10 mM Tris-HCl, pH 8, 10 mM NaCl, 0.2 % NP-40). Nuclei pellet was incubated in 0.5% SDS at 62 °C for 10 min and then quenched by water and 10% Triton X-100. MboI (NEB, R0147) enzyme digested DNA for 2 hours at 37 °C. Biotin labeled dATP (Thermo,19524016) and Klenow (NEB, M0210) were used to fill restriction fragment overhangs. The DNA was re-ligated by T4 DNA ligase (NEB, M0202) at room temperature for 4 hours. Re-ligated nuclei were lysis by 0.5% SDS and sheared for 2 min by sonication before extensive centrifugation. Supernatant was incubated at 65 °C over-night with Protease K and the DNA was purified. For libraries construction, set aside DNA by 150 ng to the biotin capture step. Biotin labeled DNA was pre-bound to Streptavidin C-1 beads (Thermo, 65-001) in biotin binding buffer. After Tween buffer and TD buffer washing, Tn5 transposition was performed on beads at 55 °C for 10 min. Libraries were generated by PCR amplification with Nextera adapters added after washing the beads. Samples were size selected by PAGE purification (300–700 bp) for effective paired end sequencing and adapter dimer removal.
GEO Accession
GSM4212721