Fig 1.

Swine PB1-F2 expression in transfected and infected cells. (A) Nucleotide phylogram comparing A/swine/Ohio/511445/2007(H1N1), A/Mexico/4108/2009(H1N1), A/swine/Nebraska/02013/2008(H1N1), A/swine/Indiana/A00968373/2012(H3N2), A/Puerto Rico/8/1934(H1N1), A/Brevig Mission/1/1918(H1N1), A/Indonesia/CDC1047/2007(H5N1), A/Vietnam/1203/2004(H5N1), and swine consensus PB1-F2 sequence. The swine consensus sequence was established using unique H1N1 and H3N2 PB1-F2 sequences collected from 2000 to the present. Bootstrap values are either shown above the branches or indicated by the dotted lines. (B) MDCK cells were transfected with pOH07/PB1-F2/3×FLAG (top row) or pOH07/PB1-F2/EGFP (bottom row). Twenty-four hours posttransfection, the cells were fixed and stained with swine PB1-F2 antibody (middle column) and anti-FLAG antibody (top left) or inherent EGFP fluorescence (bottom left). A merged image is also shown. (C) Diagram of reverse-genetics plasmid encoding the OH07 PB1-F2 in the context of the Mx09 PB1 gene. pro, promoter; term, terminator. (D) MDCK, A549, and PK-15 cells were infected with OH07 (top row), rMx/MOM (middle row), and rMx09 (bottom row). At 12 h p.i., the cells were immunostained with antibodies against NP (left columns) and OH07 PB1-F2 (right columns), followed by Alexa Fluor-conjugated secondary antibodies. Bars, 10 μm.