GSM7881750: D0_TKO_CTCF_HiChIP_rep1; Mus musculus; OTHER
Hi-ChIP experiments were performed using the Arima Hi-ChIP kit (Arima Genomics) according to the manufacturer's instructions. 15g of chromatin were used per sample and experiments were performed in duplicates. Briefly, cells were cross-linked with 2% formaldehyde for 10 min at room temperature, lysed and chromatin was digested with two different restriction enzymes included in the kit. Overhangs were filled-in in the presence of biotinylated nucleotides, followed by ligation. Ligated DNA was sonicated using the Covaris M220 to an average fragment size of 500 bp with the following parameters (Peak incident power: 50; Duty factor: 10%; Cycles per burst: 200; Treatment time: 250s). DNA was then immunoprecipitated overnight using 2.5µg of H3K27Ac (Active Motif 91193) or CTCF antibody (Active Motif 91285). After a double-size selection to retain DNA fragments between 200 and 600 bp using Ampure XP beads (Beckman Coulter) the biotin-ligated DNA was precipitated with streptavidin-coupled magnetic beads (included in the kit). The Hi-ChIP libraries were prepared on beads using the Accel-NGS 2D library Kit (Swift bioscience) following instructions from the Arima Hi-ChIP kit. Final libraries were analyzed using 4200 TapeStation system (Agilent) and sequenced. The Hi-ChIP libraries were prepared on beads using the Accel-NGS 2D library Kit (Swift bioscience) following instructions from the Arima Hi-ChIP kit. Final libraries were analyzed using 4200 TapeStation system (Agilent) and sequenced. The Hi-ChIP libraries were prepared on beads using the Accel-NGS 2D library Kit (Swift bioscience) following instructions from the Arima Hi-ChIP kit. Final libraries were analyzed using 4200 TapeStation system (Agilent) and sequenced. The Hi-ChIP libraries were prepared on beads using the Accel-NGS 2D library Kit (Swift bioscience) following instructions from the Arima Hi-ChIP kit. Final libraries were analyzed using 4200 TapeStation system (Agilent) and sequenced.