GSM3701976: ISC_ChIP_H3K9me3; Mus musculus; ChIP-Seq
GFP high CD24low ISCs were obtained from Lgr5-EGFP-IRES-creERT2;setdb1fl/fl mice (n=12) by FACS. The protocol of ChIP-seq was performed as previously described with slight emendations. Briefly, approximately 5 x 10^6 cells were cross-linked with 1% formaldehyde for 10min at room temperature, followed by quenching in 125nm glycine. Cell nuclei were obtained by lysing while cells in lysing buffer(50mM HEPES/KOH ph7.6, 1mM EDTA，140nM NaCL, 10%v/v Glycerol, 0.5% NP-40, 0.25% v/v Triton X-100) supplemented with protease inhibitor. Nuclei were subjected to sonication to acquire DNA fragments of 200-350 bp. The sonicated chromatin was applied to immunoprecipitation by incubation with H3K9me3 antibody(2ug) overnight at 4℃, followed with collected using 50ul protein A/G plus agarose beads(Millipore). Subsequent process were implement by following the protocol. The ChIP libraries were prepared using KAPA HyperPrep Kits (Roche, 7962347001)