Genotyping and screening of reassortant live-attenuated influenza B vaccine strain.
Live-attenuated influenza virus vaccines can be generated by reassortment of gene segments between an attenuated donor strain and a virulent wild-type virus. The annual production schedule for the seasonal influenza vaccine necessitates rapid and efficient genotyping of the reassorted progeny to identify the desired vaccine strains. This study describes a multiplex RT-PCR system capable of identifying each gene segment from the cold-adapted attenuated donor virus, B/Lee/40ca. The specificity of the amplification system was optimized by testing various wild-type influenza B viruses. The resulting RT-PCR method is sensitive and efficient enough for routine identification of reassortant clones to identify the desired gene constellation, consisting of six segments from the attenuated donor virus and the H and N genes from the wild-type virus. By providing a more rapid and efficient means of genotyping the candidate reassortant strains, this method could be implemented to expedite the generation of each component strain and allow more time to culture and process the final seasonal influenza vaccine.
Lee
Eun-Young
EY
Department of Biotechnology, College of Life Science and Biotechnology, Yonsei University, Seodaemun-Gu, Seoul 120-749, South Korea.
Lee
Kwang-Hee
KH
Jung
Eun-Ju
EJ
Jang
Yo Han
YH
Seo
Sang-Uk
SU
Kim
Hyun-Ah
HA
Seong
Baik Lin
BL
D003080:Cold Temperature; D017931:DNA Primers; D018076:DNA, Complementary / Q000235:genetics; D006801:Humans; D009981:Influenza B virus / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D007252:Influenza Vaccines / Q000235:genetics*; D007251:Influenza, Human / Q000517:prevention & control* / Q000821:virology; D016865:Reassortant Viruses / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D014613:Vaccines, Attenuated / Q000235:genetics; D014764:Viral Proteins / Q000235:genetics
D017931:DNA Primers; D018076:DNA, Complementary; D007252:Influenza Vaccines; D014613:Vaccines, Attenuated; D014764:Viral Proteins