Fig. 1. Characterization in tissue culture of synthetic codon pair-deoptimized Influenza viruses.

(A) Plaque phenotypes on MDCK cells of PR8 wild type virus and synthetic PR8 derivatives, carrying one (NP^Min, HA^Min, PB1^Min), two (NP/HA^Min; HA/PB1^Min) or three (PR8^3F) deoptimized gene segments. (B) Growth kinetics of PR8 wild type virus and three synthetic PR8 derivatives in MDCK cells after infection with 0.001 MOI of the indicated viruses. All combinations grew within log 8-9 although only PR8-HA^Min, PR8-NP/HA^Min and PR8^3F are shown as compared to wild type. (C) Analysis of Influenza virus protein expression in infected cells. MDCK cells were infected with PR8 wild type virus or synthetic PR8 derivatives, carrying one deoptimized gene segment each (NP^Min, HA^Min, PB1^Min), as indicated. Western blot (WB) analysis of proteins extracted from whole cell lysates was carried out with PB1, NP, HA, or actin antibodies. Actin was used to indicate equal protein loading.