GSM5013422: Mock infected pCHi-C replicate 1; Homo sapiens; Hi-C
Crosslinking was performed with 2% formaldehyde followed by cell membrane lysis. Lysates were snap frozen and stored at -80C. PCHi-C (Schoenfelder et al., 2015) was combined with Hi-C library generation as described previously (Nagano et al., 2017), with some modifications. Cell lysates were digested with DpnII (NEB) overnight at 37C rotating (950rpm). Restriction overhangs were filled-in with Klenow (NEB) using biotin-14-dATP (Jena Bioscience), and ligation was performed in ligation buffer for 4 hours at 16C (T4 DNA ligase; Life Technologies). After overnight decrosslinking at 65C, the ligated DNA was tagmented to produce fragments of 300-700 bp range. Ligation products were isolated using MyOne C1 streptavidin beads (Life Technologies), followed by washing with Wash&Binding buffer and nuclease-free water. Isolated Hi-C ligation products on the beads were then used directly for PCR amplification, and the final Hi-C library was purified with AMPure XP beads (Beckman Coulter). Promoter Capture Hi-C was performed using a custom-design Agilent SureSelect system following the manufacturer's protocol.