title: Single-reaction genomic amplification accelerates sequencing and vaccine production for classical and Swine origin human influenza a viruses.
abstract: Pandemic influenza A viruses that emerge from animal reservoirs are inevitable. Therefore, rapid genomic analysis and creation of vaccines are vital. We developed a multisegment reverse transcription-PCR (M-RTPCR) approach that simultaneously amplifies eight genomic RNA segments, irrespective of virus subtype. M-RTPCR amplicons can be used for high-throughput sequencing and/or cloned into modified reverse-genetics plasmids via regions of sequence identity. We used these procedures to rescue a contemporary H3N2 virus and a swine origin H1N1 virus directly from human swab specimens. Together, M-RTPCR and the modified reverse-genetics plasmids that we designed streamline the creation of vaccine seed stocks (9 to 12 days).
authors: Wadsworth Center, New York State Department of Health, Albany, New York 12201, USA.
authors: Bin
authors: B
authors: Zhou
authors: Matthew E
authors: ME
authors: Donnelly
authors: Derek T
authors: DT
authors: Scholes
authors: Kirsten
authors: K
authors: St George
authors: Masato
authors: M
authors: Hatta
authors: Yoshihiro
authors: Y
authors: Kawaoka
authors: David E
authors: DE
authors: Wentworth
mesh_terms: D000818:Animals; D001483:Base Sequence; D023281:Genomics; D053118:Influenza A Virus, H1N1 Subtype / Q000235:genetics*; D053122:Influenza A Virus, H3N2 Subtype / Q000235:genetics*; D009980:Influenza A virus / Q000235:genetics*; D007252:Influenza Vaccines / Q000235:genetics*; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D012367:RNA, Viral / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D013552:Swine
chemical_list: D007252:Influenza Vaccines; D012367:RNA, Viral