Cell culture, RNAi experiments, and adenoviral infection

Primary myoblasts were plated on collagen-coated plates fed with growth medium (GM) (Ham’s F-10 medium supplemented with 20% FBS and 2.5 ng/mL bFGF). Until cells were confluent to approximately 70-80% of plate, primary myoblasts can be infected with adenovirus overexpressing Cre or control virus particles re-suspended in GM at appropriate titer. After 12 hours of infection, cells were washed with DPBS warmed to 37°C to remove virus containing medium. Then cells were switched to differentiation medium (DM) (Dulbecco’s modified Eagle’s medium with 2% horse serum) to differentiate into myotubes. After 72 hours of incubation, myotubes were harvested to further investigation. The medium was changed daily.