GSM7873874: Hi-C_hst2d; Saccharomyces cerevisiae; Hi-C
Cell fixation with 3% formaldehyde (Sigma-Aldrich, Cat. F8775) was performed as described in Dauban et al. (2020). Quenching of formaldehyde with 300 mM glycine was performed at 4°C for 20 min. Hi-C experiments were performed with a Hi-C kit (Arima Genomics) with a double DpnII + HinfI restriction digestion following manufacturer instructions. Briefly, Cells were resuspended in 500ul water and transferred in VK05-0.5ml (ozyme, P000934-LYSK0-A.0) tubes containing glass beads before mechanical cells lysis. 8ul of samples were permeabilized by sequentially adding lysis buffer (15min at 4C), conditioning solution (10min at 62C) and stop solution 2 (15min at 37C) to the samples. DNA was digested using a mix of buffer A, DpnII, and Hinf1 (45min at 37C followed by 20min at 65C). DNA was repaired and biotin was added by adding a mix of buffer B and enzyme B for 45min at room temperature. DNA was re-ligated by adding a mix of buffer C and enzyme C during 15min at room temperature. Samples were then digested by protease and de-crosslinked by adding a mix of buffer D, enzyme D and buffer E during 30 min à 55°C followed by 90 min at 68°C. Samples were purified using AMPure XP beads (Beckman A63882), recovered in 120ul H2O and sonicated using Covaris (DNA 300bp) in Covaris microTUBE (Covaris, 520045). Biotinylated DNA was loaded on Dynabeads™ Streptavidin C1 (FISHER SCIENTIFIC, 10202333). Preparation of the samples for paired-end sequencing on an Illumina NextSeq500 (2x35 bp) was performed using Invitrogen TM Collibri TM PS DNA Library Prep Kit for Illumina and following manufacturer instructions.