HiChIP of Undifferentiated LUHMES 3A
Technical replicates for LUHMES neurons and undifferentiated cells were harvested and flash frozen.  Chromatin was fixed with disuccinimidyl glutarate (DSG) and formaldehyde in the nucleus. Fixed chromatin was digested in situ with micrococcal nuclease (MNase) and then extracted upon cell lysis. Chromatin fragments were incubated with the CTCF antibody overnight for chromatin immunoprecipitation. The antibody-protein-DNA complex was pulled down with protein A/G-coated beads. Chromatin ends were repaired and ligated to a biotinylated bridge adapter followed by proximity ligation of adapter-containing ends. After proximity ligation, crosslinks were reversed, the DNA was purified from proteins and converted into a sequencing library. The sequencing library was generated using Illumina-compatible adapters. Biotin-containing fragments were isolated using streptavidin beads before PCR enrichment of the library.