Full length sequencing of all nine subtypes of the neuraminidase gene of influenza A viruses using subtype specific primer sets.
An RT-PCR based method was developed using subtype specific overlapping primers to obtain full length amplification of neuraminidase (NA) gene from all subtypes (N1-N9) of influenza A viruses. This method was validated using reference strains of avian influenza viruses (AIV) (N1-N9), human influenza viruses (N1 and N2), and swine influenza viruses (N1-N3). Amplification of the NA gene was obtained with all viruses tested. Additionally, 200 field isolates of AIV from wild birds were tested by this method and the NA gene was amplified in all isolates. The NA subtype of all 200 isolates was determined by further sequencing of the amplified NA genes and all sequences were submitted to GenBank. The method described in this paper can be used to determine subtype of influenza isolates as well as their evolution and mutations if any, in the NA gene.
Chander
Yogesh
Y
Department of Veterinary Population Medicine, College of Veterinary Medicine, University of Minnesota, 1333 Gortner Avenue, Saint Paul, MN 55108, United States.
Jindal
Naresh
N
Stallknecht
David E
DE
Sreevatsan
Srinand
S
Goyal
Sagar M
SM
D000818:Animals; D001717:Birds; D017931:DNA Primers / Q000235:genetics*; D005838:Genotype; D006801:Humans; D009980:Influenza A virus / Q000235:genetics* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D009439:Neuraminidase / Q000235:genetics*; D012367:RNA, Viral / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D017422:Sequence Analysis, DNA; D013552:Swine; D014764:Viral Proteins / Q000235:genetics*
D017931:DNA Primers; D012367:RNA, Viral; D014764:Viral Proteins; C487630:NA protein, influenza A virus; D009439:Neuraminidase