Figure 1

Construction and characterization of the siDEN suppressor. (A) Diagram of the construction of the plasmid vector, pSMWZ-1, capable of expressing a DEN infection suppressor cassette. Abbreviations: N*, Not I; K, Kpn I; A, Apa I; E, EcoR I; SUP-1, Suppressor cassette; (B) Co-transfection with pSMWZ-siEGFP and pEGFP inhibits the expression of EGFP in cultured cells. HEK293 cells were transfected with different concentrations of plasmid DNA and three days later, EGFP-positive cells were counted by fluorescence microscopy. Results are expressed as mean ± SEM. *p < 0.05 compared to control (siRSVNS1, an unrelated siRNA construct against respiratory syncytial virus). (C) pSMWZ-siDEN suppression of DEN-2 virus replication. Vero cells were transfected with pSMWZ-siDEN3UT or pSMWZ-siDENpreM plasmid. Two days later, the cells were infected with DEN-2 virus (MOI of 0.1) and 5 days later, the numbers of DEN-2 virus infected cells were counted by fluorescence microscopy. Data are mean ± SEM. *p < 0.05 compared to control DEN-2. (D) AAV-siEGFP inhibits the expression of EGFP in cultured cells. HEK293 cells were infected with different concentrations of AAV-siEGFP, and three days later the cells were transfected with pEGFP. EGFP-positive cells were counted by fluorescence microscopy. Statistically significant differences, **p < 0.01, when compared to pEGFP plasmid control, AAV-siRSV (10⁷) and AAV-siEGFP (10⁸) group, respectively.