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#Transfer cells to serum **and P/S/G free** media
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#Split 293A cells into 10-150 mm dishes in DMEM/FBS with '''no antibiotics'''.
#Transfect when cells are 90-95% confluent.
#Combine 500 ug DNA with 10 mL OptiMEM, and separately 700 uL Lipofectamine 2000 with 10 mL OptiMEM
#After 5 minutes, combine the DNA solution with the Lipofectamine solution.
#Treat cells for 30min with 100 nM Wortmannin to suppress endogenous S6K phosphorylation
#Wash cells with D-PBS -/- twice (10 mL per plate)
#Add 0.5 mL [[HNTG Buffer]] per to each plate and scrape
#Collect scraped cells and incubate end over end in eppendorf tube for 30 min
#Centrifuge in eppendorf centrifuge for 10 min at 14 000 RPM