GSM9410355: 22Rv1_RCMC_NTC_R2; Homo sapiens; OTHER
For RCMC, cells were fixed with 3mM DSG and 1% formaldehyde. After quenching, cells were lysed on ice to obtain nuclei. Chromatin was digested in situ with MNase to fragment the DNA at nucleosome resolution. DNA was repaired with biotinylated dNTPs, and then free DNA ends were proximity ligated while unligated ends were digested. Chromatin was then reverse crosslinked and digested by Protease K. Lastly, DNA was purified by phenol chlroroform extraction and purified. 250 - 400 bp dinucleosomal DNA fragments were selected on an agarose gel and end polished. Streptavidin beads were used to pull down ligated dinucleosomal DNA fragments. RCMC library: DNA fragments bound to streptavidin beads were made into libraries using NEBNext Ultra II DNA Library Prep Kit (NEB) nad KAPA HiFi HotStart ReadyMix (Roche)following manufacturer's instructions. Purified libraies were pooled together and made into RCMC libaries following the Twist Standard Hybridization Target Enrichment Protocol (Twist Biosciences). Briefly, the libraries were hybridized to custom probe panels for loci of interest on chromosome 6 and bound to streptavidin beads. DNA fragments on the beads were amplified using Equinox Library Amp Mix (Twist Biosciences) and purified.