Effect of siRNAs on the expression of HBV surface open reading frame in HEK293 and T98G cells. (A) As shown in Additional file 2: Figure S2. (B) Fluorescence micrographs of cells transfected with reporter plasmids and cotransfected with either the corresponding or non-corresponding siRNA with Lipofectamine TM 2000 (Invitrogen). At 24 hrs after transfection, the cells were observed with an Olympus BH-2 microscope, and representative bright-field images (left column) and relative fluorescent-field images (right column) were recorded by tenfold amplification. (C) Flow cytometry analysis of siRNA-mediated gene silencing of EGFP. Level of EGFP expression in cells cotransfected with (a) pEGFP-N1 vector; ( b) pS-EGFP-N1 and pU6; (c) pS-EGFP-N1 and S1; (d) pS-EGFP-N1 and S2; (e) pS-EGFP-N1 and S3 (heterologous siRNA). (D) Cells were analyzed for EGFP expression by fluorescence-activated cell sorting and the level of fluorescence relative to the control was quantitated. The mean fluorescence intensity of control siRNA was taken as 100% and adopted as control. Data represent means±SD from three independent experiments were performed in triplicate.*Significant differences compared to the heterologous siRNA control: Student’s t-test; P < 0.05.