title: GSM8791661: Mouse MuSC DSC G4 bio rep2; Mus musculus; ChIP-Seq
library_construction_protocol: Skeletal muscle tissue from mouse hindlimb and forelimb muscles were dissected, gently minced with blades and digested with Collagenase II (1000U/ml) in Ham's F10 for 90 minutes in a shaking water bath at 37°C. Digested tissue was then washed twice with rinsing media (10% HS, in Ham's F-10) and centrifuged at 700 xg at 4°C for 5 minutes. Second digestion was performed by adding Collagenase II (1000U/ml) and Dispase (11U/ml) in rinsing media and incubated in shaking water bath at 37°C for 30 minutes. Digested tissue was passed through a 20-gauge needle for 12 times and filtered through a 40 μm filter followed by spinning at 700 xg for 5 min at 4°C. For mouse muscles, MuSCs were then sorted out by FACS Aria Fusion (BD) and collected as GFP+ groups. For human muscles, cells were stained with antibodies and FITC-(CD45-, CD31-, CD34-) APC+(CD29+) PE-CY7+ (CD56+) cells were sorted out as human MuSCs. Isolated satellite cells were immediately collected or were cultured in Ham's F10 supplemented with 20% FBS and bFGF (0.025 μg/ml) (growth medium) for other assays. CUT&RUN protocol is referred to CUT&RUN assay kit (Cell Signaling Technology, 86652,). For DNA library construction, a NEBNext® Ultra™ II DNA Library Prep Kit for Illumina® (NEB, E7645S) was used according to the manufacturer's instructions.