title: A reverse transcription-PCR for subtyping of the neuraminidase of avian influenza viruses.
abstract: To date, nine neuraminidase (NA) subtypes of avian influenza viruses have been identified. In order to differentiate the NA of avian influenza viruses rapidly, a reverse transcription PCR (RT-PCR) was developed. Nine pairs of NA-specific primers for the RT-PCR were designed based on the analysis of 509 complete NA sequences in GenBank. The primers were designed to amplify partial NA genes and each pair is unique to a single NA subtype (N1-N9). By nine RT-PCRs simultaneously in a set of separate tubes, the subtype of NA was determined by subsequent agarose gel electrophoresis and ethidium bromide staining, since only one of the nine RT-PCRs would give a product of expected size for each virus strain. In comparison with the established method of sequence analysis of 101 reference strains or isolates of avian influenza viruses, the RT-PCR method had a sensitivity of 97.3% and a specificity of 91.1% in subtyping avian influenza viruses. These results indicate that the RT-PCR method described below provides a specific and sensitive alternative to conventional NA-subtyping methods.
authors.affiliation: Animal Infectious Disease Laboratory, School of Veterinary Medicine, Yangzhou University, Yangzhou, Jiangsu 225009, China.
authors.forename: Bao-Feng
authors.initials: BF
authors.lastname: Qiu
authors.forename: Wu-Jie
authors.initials: WJ
authors.lastname: Liu
authors.forename: Da-Xin
authors.initials: DX
authors.lastname: Peng
authors.forename: Shun-Lin
authors.initials: SL
authors.lastname: Hu
authors.forename: Ying-Hua
authors.initials: YH
authors.lastname: Tang
authors.forename: Xiu-Fan
authors.initials: XF
authors.lastname: Liu
mesh_terms: D000818:Animals; D017931:DNA Primers; D004372:Ducks / Q000821:virology*; D009980:Influenza A virus / Q000145:classification* / Q000201:enzymology* / Q000235:genetics; D009439:Neuraminidase / Q000145:classification* / Q000235:genetics*; D012367:RNA, Viral / Q000235:genetics; D012015:Reference Standards; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity
chemical_list: D017931:DNA Primers; D012367:RNA, Viral; D009439:Neuraminidase