FIG. 4.

Level of viral plus-strand RNAs produced in siRNA-transfected cells. MA104 cells were transfected with the indicated siRNA and infected with SA11-5N or DxRRV. (A) After cells were maintained in [³²P]orthophosphate from 3 to 9 h p.i., RNAs were recovered and analyzed by electrophoresis on a urea-polyacrylamide gel and by autoradiography. The positions of viral plus-strand RNAs (1 to 11) and dsRNAs (ds1 to ds9) are shown. (B and C) Unlabeled RNAs were recovered from cells at 6 h p.i., resolved by electrophoresis, and transferred to nylon membranes. Blots were probed with ³²P-labeled RNAs specific for SA11 g5 (SA11-g5PR) or g8 (SA11-g8PR) or for DxRRV g4 (RRV-g4PR) or g9 (D-g9PR), and the intensity of the bands was determined with a phosphorimager. The levels of the g9 and g5 plus- strand RNAs were calculated by dividing their band intensities by the corresponding intensities determined for g4 or g8 plus-strand RNAs, respectively. The values were then normalized, with the value for the IR samples taken as 100%.