title: GSM3371604: BMHSC_iHiC_Rep2; Mus musculus; Hi-C
library_construction_protocol: E14.5 fetal liver and adult bone marrow were dissected and HSCs were purified using cell surface marker: Lin-Sca1+cKit+CD135-. Half million sorted cells were cross-linked with 1% formaldehyde for 10 min at RT and quenched with glycine (final concentration 0.125M). Nuclei were then permeabilized with 250 μL of cold lysis buffer (10 mM Tris, 10 mM NaCl, 0.2% Igepal CA630) and 50 μL of protease inhibitors (P8340, Sigma). Chromatin was digested with 100 units of MboI (R0147, NEB) overnight at 37°C with rotation. Restriction fragment ends were labelled with biotinylated nucleotides and proximity ligation in a small volume with 5 μL of 400 U/μL T4 DNA ligase (B0202, NEB). After reversal of cross-link, DNA was sheared to a length of 300~500 bp and size-selected using AMPure XP beads (A63881, Beckman Coulter). The ligated junctions were pulled down with 150 μL of Dynabeads MyOne Streptavidin C1 magnetic beads (65001, Thermo Scientific). Libraries were constructed using NEB DNA quick ligase (NEB, M2200) and customized Illumina indexed adapters. Libraries were size-selected at 200-500 bp by AMPure XP beads. Libraries were quantified with KAPA qPCR and bioanalyzer prior to single-end sequencing.
attributes.tag: GEO Accession
attributes.value: GSM3371604