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Generation of recombinant influenza virus by reverse genetics.
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The pHW2000 eight-plasmid system (26) was adopted to generate desired recombinant viruses after whole-genome sequencing. Mutations were introduced into desired genes on pHW2000 plasmids via site-directed mutagenesis. Recombinant plasmids were transfected into cocultured 293T and MDCK cells with Lipofectamine 2000 (Life Technologies) according to the manufacturer's instructions. The infectious particles from supernatant were harvested at 72 h posttransfection, and the recombinant virus titer was determined by plaque assay.
Copyright and License information: ©2016, American Society for Microbiology. All Rights Reserved.
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