Matrigel coated culture plates

C2C12 cells were cultured in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% FBS (Hyclone) and 1% penicillin, in a 37°C humidified incubator with 5% CO₂. For differentiation, C2C12 cells were plated on the BD Matrigel‐coated culture plates and differentiated in DMEM supplemented with 2% horse serum, 400 nM insulin, and 1% penicillin. Plasmids and lipofectamine 2000 mixture were transfected in C2C12, based on the manufacturer's instructions, and harvested after 24 h. For transfection, pcDNA‐GFP‐PRMT5 plasmid and pcDNA3.1‐2xFLAG‐SREBP1a plasmid were mixed with lipofectamine 2000 Reagent in Opti‐MEM media (Giobco) and transfected into 60% confluent cells following the protocol from manufacture. Opti‐MEM media was changed to growth medium after 4 h and cells were harvested after 24 h for further analysis.

Primary myoblasts were isolated from hind limb skeletal muscles of 4‐week‐old mice as previously described (Kim et al, 2020 ). Muscle tissues were minced and digested in type II collagenase and dispase B mixture (Roche). Digestion was neutralized by adding growth media containing F‐10 Ham's medium (ThermoFisher Scientific), 20% fetal bovine serum (FBS), 1% penicillin, 4 ng/ml basic fibroblast growth factor (ThermoFisher Scientific), and cells were cultured on collagen‐coated plates. Preplating was performed to purify primary myoblasts. For differentiation, primary myoblasts were plated on the BD Matrigel‐coated culture plates and differentiated in DMEM supplemented with 2% horse serum and 1% penicillin.