GSM3262885: MOUSE_SPERM_HiC_Rep2; Mus musculus; Hi-C
We generated in-situ Hi-C libraries using DpnII restriction enzyme following the protocol as described (Rao et al., 2014). Briefly, 10 million sperm were crosslinked with 1% formaldehyde, quenched with glycine, washed with PBS, and permeabilized to obtain intact nuclei. Nuclear DNA was then digested with DpnII, the 5'-overhangs were filled with biotinylated dCTPs and dA/dT/dGTPs to make blunt-end fragments, which were then ligated, reverse-crosslinked, and purified by standard DNA ethanol precipitation. Purified DNA was sonicated to 200-500 bp small fragments and captured with streptavidin beads. Standard Illumina TruSeq library preparation steps including end-repairing, A-tailing, and ligation with universal adaptors were all performed on beads, washing twice in TWB between each step. DNA on the beads was PCR amplified with barcoded primers using KAPA SYBR FAST qPCR Master Mix for 5~12 PCR cycles to obtain enough DNA for sequencing. Generated libraries were paired-end sequenced on Illumina HiSeq2500 v4. Two sets of biological replicates were generated, and replicates were combined after ensuring high correlation.
GEO Accession
GSM3262885