P-GSE45414-2 - P-GSE45414-2
sample treatment protocol
For knock down experiments, A673 cells were grown to 30-35% confluence in 100 mm dishes at which point growth medium was removed and 5 ml fresh medium added along with 5 ml of virus and Polybrene at a final concentration of 8 μg/ml. After 24 hours, medium was replaced with 2 ml fresh growth medium containing puromycin at 4 μg/ml. Cells were selected for 48 hours, then harvested for Western-blot and RNA extraction. For drug treatment experiments, A673 cells were grown to 65% confluence and then were either left untreated or treated for 24hrs with DMS0 or actinomycin D at low- (5nM) or high-dose (50nM).