FIG. 1.

Functional analysis of the influenza virus reporter amplicon with a canine polymerase I promoter in MDCK cells. (A) The negative-sense Renilla luciferase [R-LUC(−)] coding region is flanked by modified noncoding regions (NCR; hatched boxes) from the nucleoprotein gene of A/WSN/33 virus (24, 30). The sequences of the canine POL-I promoter and the canine Pol-I terminator (k9POL-I and k9TI, respectively; gray box) were fused upstream to the 5′ NCR or downstream to the 3′ NCR, respectively. (B) MDCK cells were cotransfected with reporter plasmid pk9POLI-RLuc and influenza virus RNP-expressing plasmids or control plasmid. RLU, relative light units. (C) MDCK cells transfected with pk9POLI-RLuc plasmid for 24 h were infected with influenza viruses at an MOI of 0.001. Luciferase activities in whole-cell lysates collected at 24 h postinfection are shown as the average of luciferase activity of cells from three independent wells. The values shown are the Renilla firefly activities from 10⁴ cells after normalization using firefly luciferase expression from a cotransfected plasmid to account for variation in transfection efficiency. Error bars depict standard errors, and brackets denote P values from Student's t test: *, P > 0.9; **, P < 0.05; and ***, P < 0.005.