A multiplex RT-PCR method for screening of reassortant live influenza vaccine virus strains

Suk Hoon Ha, Hyun Ah Kim, Yeon Hee Kim, Jae Seoung Kim, Kwang Hee Lee, Song Yong Park, Wan Je Park, Baik Lin Seong

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

A system based on reverse transcription polymerase chain reaction (RT-PCR) of the RNA genome was established to identify genetic composition of influenza viruses generated by reassortment between an attenuated donor virus and virulent wild type virus. The primers were designed, by multiple sequence alignment of variable regions, specific for cold-adapted donor virus HTCA-A101, as compared to other influenza A viruses. The specificity of each primer set was confirmed and the primers were combined to perform RT-PCR in multiplex manner. The multiplex PCR was adopted to distinguish the 6:2 reassortant viruses containing six internal genome segments of attenuated donor virus and two surface antigens of virulent strain from the wild type viruses. The method allowed us to optimize of the reassorting process on a routine basis and to confirm the selection of reassortant clones efficiently. The method is suitable for analyzing the contribution of specific gene segments for growth and attenuating characteristics and for generation of live attenuated vaccine by annual reassortment.

Original languageEnglish
Pages (from-to)154-163
Number of pages10
JournalJournal of Virological Methods
Volume134
Issue number1-2
DOIs
Publication statusPublished - 2006 Jun 1

All Science Journal Classification (ASJC) codes

  • Virology

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