Multi-targeting with both DIS- and vif-shRNAs results in longer-lasting inhibition of HIV-1 genes. (A) vif-shRNA-expressing cells were challenged with HIV-1_NL-4-3 and the cells were cultured. On day 21, the cells were further challenged with CS-U6-DIS. The levels of HIV-1 gag p24 antigen were reduced significantly in DIS-shRNA-expressing cells compared to vif-shRNA cells alone for 34 days. vif-shRNA, solid square; negative control, solid diamond; vif-shRNA/DIS-shRNA, solid triangle. (B) Expression of transgenic EGFP in Sup-T1 cells expressing vector transgenes was examined by fluorescence microscopy. a, vif-shRNA at 7 days; b, vif-shRNA at 21 days; c, vif-shRNA/DIS-shRNA at 21 days; d, vif-shRNA/DIS-shRNA at 28 days; e, vif-shRNA/DIS-shRNA at 34 days.