GSM5209155: Capture-C in E11.5 distal-limbs from ΔF-all mutant embryos; Mus musculus; Hi-C
Fixed nuclei were extracted adding ice-cold custom-made lysis buffer containing the following components: 50mM Tris HCl pH 7.5, 150 mM NaCl, 5 mM EDTA, 0.5% NP-40, 1.15% Triton X-100, 25X Protease Inhibitor in Milliwater. Extracted nuclei were then collected, snap frozen and stored at -80 °C or further processed with the following steps: digestion with DpnII RE, re-ligation of the digested fragments, de-crosslinking and DNA purification (3C fragments). The 3C library was further processed using SureSelectXT Target Enrichment System for the Illumina Platform (Agilent Technology). 200 ng or 3 μg of sample was shared using Covaris sonicator and the following parameters; duty cycle: 10%, intensity: 5, cycle per burst:200, time: 6 minutes, temperature: 4°C. Shared sample was then treated following the kit guidelines (end repair, dA-tailing, adaptor ligation, PCR enrichment of Adaptor ligated DNA, DNA purification, hybridization and capture). The hybridization was performed using SureSelect XT Custom RNA probes (Cat # 5190-4836) and the capture with Streptavidin-Coated Beads (Invitrogen). PCR enrichment and sample indexing were done following Agilent instructions. Capture libraries were sequenced with 75bp paired-end reads.