For lentiviral infection in 293T or HAP1 cell lines, 2.5 × 105 cells were seeded in six-well plates or 6 × 104 cells in 24-well plates for HAP1 cells infected with the NLuc reporter plasmid. 24 hours after initial seeding, 1 ml of virus (or media for mock infection) along with 2 ml of media supplemented with 10 μg/ml of polybrene was added to each well. The cells were washed with PBS and fed fresh media 24 h post-infection. Puromycin selection begun 48 h after infection at a concentration of 2 μg/ml. Fresh media supplemented with puromycin was added every other day and continued until the mock infection had no observable living cells. Proper integration and expression of each construct was verified via immunoblotting.
