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Reverse Genetics of Newcastle Disease Virus

  • Stivalis Cardenas-Garcia
  • Claudio L. AfonsoEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1602)

Abstract

Reverse genetics allows for the generation of recombinant viruses or vectors used in functional studies, vaccine development, and gene therapy. This technique enables genetic manipulation and cloning of viral genomes, gene mutation through site-directed mutagenesis, along with gene insertion or deletion, among other studies. An in vitro infection-based system including the highly attenuated vaccinia virus Ankara strain expressing the T7 RNA polymerase from bacteriophage T7, with co-transfection of three helper plasmids and a full-length cDNA plasmid, was successfully developed to rescue genetically modified Newcastle disease viruses in 1999. In this chapter, the materials and the methods involved in rescuing Newcastle disease virus (NDV) from cDNA, utilizing site-directed mutagenesis and gene replacement techniques, are described in detail.

Key words

Newcastle disease virus Reverse genetics Virus rescue Site-directed mutagenesis Gene replacement cDNA Cell culture 

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Copyright information

© Springer Science+Business Media LLC 2017

Authors and Affiliations

  1. 1.Southeast Poultry Research LaboratoryUnited States Department of AgricultureAthensUSA
  2. 2.Department of Population Health, Poultry Diagnostic and Research Center, College of Veterinary MedicineThe University of GeorgiaAthensUSA

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