Single-reaction genomic amplification accelerates sequencing and vaccine production for classical and Swine origin human influenza a viruses.
J Virol, 2009/10;83(19):10309-13.
Zhou B[1], Donnelly ME, Scholes DT, St George K, Hatta M, Kawaoka Y, Wentworth DE
Affiliations
PMID: 19605485DOI: 10.1128/JVI.01109-09
Impact factor: 6.549
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).
MeSH terms
Animals; Base Sequence; Genomics; Influenza A Virus, H1N1 Subtype; Influenza A Virus, H3N2 Subtype; Influenza A virus; Influenza Vaccines; Models, Genetic; Molecular Sequence Data; Mutation; Plasmids; RNA, Viral; Reverse Transcriptase Polymerase Chain Reaction; Sequence Analysis, DNA; Swine
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