African Journal of
Microbiology Research

  • Abbreviation: Afr. J. Microbiol. Res.
  • Language: English
  • ISSN: 1996-0808
  • DOI: 10.5897/AJMR
  • Start Year: 2007
  • Published Articles: 5233

Full Length Research Paper

Prokaryotic expression and purification of grass carp reovirus capsid protein VP7 and its vaccine potential

Yongxing He1, Qian Yang1, Hongxu Xu2, Hao Wu1, Fangyuan Wu1 and Liqun Lu1*
1Key Laboratory of Aquatic Genetic Resources and Utilization /Ministry of Agriculture, Shanghai Ocean University, 201306 Shanghai, China. 2Department of Laboratory Medicine, The First Affiliated hospital of Sun Yat-sen University, Guangzhou 510080, China.
Email: [email protected]

  •  Accepted: 10 May 2011
  •  Published: 04 July 2011

Abstract

The 11 dsRNA fragmental genome of grass carp reovirus (GCRV) is enclosed in five inner core proteins and two outer capsid proteins. The Glutathione S-transferase (GST) fusion protein expression vector pGEX-4T-3 was employed to clone and expression of GCRV outer capsid gene vp7, which was amplified by reverse transcription-polymerase chain reaction (RT - PCR) from infected Grass carp. The recombinant GST-fusion protein rVP7 was induced by 1 mM IPTG in Dh5αand confirmed by SDS-PAGE and Western blot assays using both anti-GST-tag and anti-VP7 monoclonal antisera. An expected 52-kDarVP7 was highly expressed, and was mainly exhibited in the formation of the inclusion body. After purification, rVP7 was intraperitoneally injected to the experimental mice to produce anti-rVP7 polyclonal serum. In vitro microneutralization assay indicated that polyclonal antibody against rVP7 could neutralize GCRV, and suggested that rVP7 had the potential to be used as subunit vaccine against GCRV infection. The present study paved the way for further characterization of the immunogenicity of viral outer capsid protein VP7 in grass carp Ctenopharyngodon idellus and could be based to develop antibody or antigen detection assays for GCRV pathogen.

 

Key words: Grass carp reovirus, VP7 protein, prokaryotic expression, western blot, microneutralization.