FIG. 4.

Reactivation of silenced vectors. NIL-2 cells were infected with silencing-prone vectors RNIG and RNIG3-MIE with mutated Sp1 binding sites. The cells were selected with G418, and after 6 days of selection, the antibiotic was removed. After an 18-day cultivation, the GFP-negative cells were sorted by FACS. Each GFP-negative population was then divided into two subpopulations 6 days after the sorting: one was treated with 5-AzaC and/or TSA for 4 days, and the proportion of GFP-expressing cells was assessed. The nontreated GFP-negative cell population was cultivated further, and the treatment was repeated at regular intervals. Each experiment was done in triplicate.