title: GSM4808652: WT_left_ventricle_control, rep1; Mus musculus; OTHER
library_construction_protocol: 4C templates were prepared as previously described (de Krijger et al., 2020). In short, snap frozen adult left ventricle were crushed, dissociated and dounce homogenized to obtain single cell suspension. Chromatin was cross-linked with 2% formaldehyde in PBS with 10% FCS for 10 minutes at room temperature. Nuclei were isolated and cross-linked DNA was digested with DpnII, followed by proximity ligation. Cross-links were removed and a secondary restriction digestion with Csp6I performed, followed again by proximity ligation. Per viewpoint, 8 PCR reactions were performed on 200 ng of 4C template each, to a total of 1.6 µg. PCR reactions were pooled and purified for next generation sequencing. The 4C-seq PCRs were essentially performed as previously described (de Krijfer et al., 2020) using reading primers (Nppa promoter – GCCCTAGGAGAGCTATGATC) as 4C viewpoints combined with non-reading primer ( Nppa promoter –TAACAGGGACATCTAGGGTG). The primers were extended with Illumina adapter sequences and the samples were sequenced on the MiSeq.