GSM4630462: HiC_Late_rep2; Mus musculus; Hi-C
Hippocampal tissue was extracted and flash frozen. After extraction, samples were immediately homogenized in 0.5 mL ice cold PBS with protease inhibitors (Pi) (11836170001, Roche). The suspension was fixed with 1% paraformaldehyde for 10 min, quenched with Glycine 2.5M for 5 min and washed twice with 5 mL of PBS. The suspension was centrifuged at 1200 g, 4°C, for 5 minutes and the pellet was resuspended in 5 mL NF-1 hypotonic buffer (0.5% Triton X-100/0.1M Sucrose/5mM MgCl2/1mM EDTA/10mM Tris-HCl, pH 8.0, Pi). Pelleted nuclei were resuspended in and incubated with NeuN and GFP primary antibodies for overnight in 4°C. Unbound antibodies were washed out twice and nuclei were filtered with 40 µm mesh filter for FACS sorting. Nuclei from fixed cells were sorted into 1.5 mL eppendorf tubes containing 500 µl fresh ice-cold lysis buffer (10mM Tris pH=8, 10mM NaCl, 0.2% Igepal CA-630, Pi and PCR grade water) and frozen on dry ice and stored at -80c until they were used. Hi-C libraries were prepared from 100,000 cells/per group (collected from two animals) in two biological replicates, using DovetailTM Hi-C kit manual (v.1.03, Dovetail Genomics, Chicago, USA).
GEO Accession
GSM4630462