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Reverse Genetics System for the Avian Coronavirus Infectious Bronchitis Virus

  • Erica BickertonEmail author
  • Sarah M. Keep
  • Paul Britton
Part of the Methods in Molecular Biology book series (MIMB, volume 1602)

Abstract

We have developed a reverse genetics system for the avian coronavirus infectious bronchitis virus (IBV) in which a full-length cDNA corresponding to the IBV genome is inserted into the vaccinia virus genome under the control of a T7 promoter sequence. Vaccinia virus as a vector for the full-length IBV cDNA has the advantage that modifications can be introduced into the IBV cDNA using homologous recombination, a method frequently used to insert and delete sequences from the vaccinia virus genome. Here, we describe the use of transient dominant selection as a method for introducing modifications into the IBV cDNA that has been successfully used for the substitution of specific nucleotides, deletion of genomic regions, and exchange of complete genes. Infectious recombinant IBVs are generated in situ following the transfection of vaccinia virus DNA, containing the modified IBV cDNA, into cells infected with a recombinant fowlpox virus expressing T7 DNA-dependant RNA polymerase.

Key words

Transient dominant selection (TDS) Vaccinia virus Infectious bronchitis virus (IBV) Coronavirus Avian Reverse genetics Nidovirus Fowlpox virus T7 RNA polymerase 

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Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  1. 1.The Pirbright InstitutePirbrightUK

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