GSM9196970: Ctcf_0H_4C-seq_rep2; Mus musculus; OTHER
10 million cells were collected per sample. Cells were crosslinked, lysed, and chromatin was digested with the primary restriction enzyme DpnII. After ligation, crosslinks were reversed and DNA was purified according to established 4C-seq protocols (van de Werken et al., 2012; Haarhuis et al., 2017). Purified 4C DNA was digested with the secondary restriction enzyme Csp6I and religated to generate small circular DNA fragments. These fragments were amplified by PCR using viewpoint-specific primers. Libraries were indexed using a two-step PCR method to enable multiplexed sequencing. For allele-resolved 4C-seq, a reverse primer specific to a single nucleotide variant was used. Final PCR products were purified and quantified prior to sequencing.