Development of a reverse genetics system based on RNA polymerase II for Newcastle disease virus genotype VII

Virus Genes. 2015 Feb;50(1):152-5. doi: 10.1007/s11262-014-1137-x. Epub 2014 Nov 11.

Abstract

Newcastle disease virus (NDV) has only a single serotype but diversified genotypes. Genotype VII strains are the prevalent currently circulating genotype worldwide, and in particular, these strains cause outbreaks in waterfowl. In this study, a reverse genetics system for highly virulent NDV isolated from goose flocks was developed independent of conventional T7 RNA polymerase. Infectious virus was successfully generated by an RNA polymerase II promoter to drive transcription of the full-length virus antigenome. A green fluorescent protein (GFP)-expressing virus was generated by inserting an additional transcription cassette coding for the enhanced GFP between the P and M genes of the genome. The expression of GFP was confirmed by western blotting and fluorescence microscopy. The replication kinetics and pathogenicity of the recombinant viruses are indistinguishable from the parental wild-type virus. This reverse genetics system will provide a powerful tool for the analysis of goose-origin NDV dissemination and pathogenesis, as well as preparation for genotype-matched NDV attenuated vaccines.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Artificial Gene Fusion
  • Blotting, Western
  • Geese / virology
  • Genes, Reporter
  • Genome, Viral
  • Genotype
  • Green Fluorescent Proteins / analysis
  • Green Fluorescent Proteins / genetics
  • Microscopy, Fluorescence
  • Newcastle disease virus / genetics*
  • Newcastle disease virus / isolation & purification
  • Newcastle disease virus / pathogenicity
  • Newcastle disease virus / physiology
  • Promoter Regions, Genetic
  • RNA Polymerase II / genetics
  • RNA Polymerase II / metabolism*
  • Reverse Genetics / methods*
  • Transcription, Genetic
  • Virology / methods*
  • Virus Replication

Substances

  • Green Fluorescent Proteins
  • RNA Polymerase II