Viral RNA polymerase activity and Viral growth kinetics in MDCK and A549 cells.

(A) Polymerase activity of SD16 with different PB2 mutations in a minigenome assay. Four protein expression plasmids (PB2, PB1, PA, NP) for the RNP combinations were transfected into 293T cells together with luciferase reporter plasmid pYH-Luci and internal control plasmid Renilla, as described in Materials and Methods. The values shown are means ± SD of results for three independent experiments and are standardized to the activity of SD16 (100%). *, P<0.05 compared with that of SD16-infected cells. (B, C, D, E) Viral growth kinetics of reassortant viruses and PB2 mutants in MDCK and A549 cells. Confluent momolayers of cells were infected with rescued H9N2 viruses at an MOI of 0.01. Cell supernatants were harvested every 12 hours until 60 hpi and titrated by plaque assay on MDCK cells. Each data point represents the mean virus yield from three individually infected wells ± SD. (B) reassortant viruses in MDCK cells, (C) PB2 mutants in MDCK cells, (D) reassortant viruses in A549 cells, (E) PB2 mutants in A549 cells.