Plasmid DNA of the control pGIPZ lentiviral vectors (with Puromycin selection and TurboGFP mark) or with survivin shRNA (V2LHS_262484) [22] were first prepared. HEK 293T packaging
Abiraterone acetate were then seeded in a 100 mm cell cultural dish at 80% confluence and incubated for 24 h at 37 °C and 5% C02. Then 250 μl DMEM containing 2.5 μg pGIPZ survivin shRNA (or empty pGIPZ vector), 2.5 μg psPAX2 (or pCMV-dR8.74), 1.0 μg pMD2.G in one tube mixed with 250 μl DMEM containing 9–12 μl lipofectamine and stayed at room temperature for 20 min. The cell medium in the 293T cell dish was gently replaced with 500ul DNA/Lipo complex. Three ml 293T media (DMEM with 10%FBS and 1% Pen/Strep) were added after a few minutes at room temperature for 16 h at 37 °C and 5% CO2. The medium in the dish was replaced with new 293T media next day, and the dish was incubated for additional 24 h at 37 °C with 5% CO2. Virus-containing supernatant was harvested and filtered through 0.45 μm cellulose acetate syringe filter, and store virus at 4 °C. Check TurboGFP expression before collection of virus in the supernatant. The dish with 293T
cells was added another 3.0–3.5 ml 293T media and incubated for overnight at 37 °C with 5% CO2. The supernatant was collected as above and combined together and stored at 4 °C for target cell infection. The 293T cell dish was discarded or were check for gene expression.
