FIG 7.

PB2 and PA mutational effects on viral polymerase activity by minigenome assay. 293T cells were transfected with plasmids encoding PA, PB1, PB2, and NP, together with pPolI-358Luc and pβ-gal. Wild-type PB1 and NP plasmids from A/TN/1-560/09 were used in each group, while PB2 and PA plasmids carrying mutants identified in MA9-22 were varied. (A and B) After 24-h incubation at 33°C (A) and 37°C (B), both luciferase and β-Gal expression levels were measured. β-Gal expression was used to normalize the data. R.L.U., relative light units. (C) PB2 mutants carrying nonsynonymous and/or synonymous mutations from MA9-22 were compared at 37°C. The reported values are means and standard deviations from 3 independent experiments (n = 9). *, P < 0.05; **, P < 0.01; ***, P < 0.001.