J Vet Sci.  2002 Mar;3(1):13-18.

Expression of Open Reading Frame 5 Protein of Porcine Reproductive and Respiratory Syndrome Virus Using Semliki Forest Virus Expression System

• Jung HS
• Hwang IW
• Kim SM
• Kim CJ
• Shin KS
• Kim HS

Abstract

The ORF5 gene encodes a major envelope glycoprotein (GP5), which is one of the three major proteins of porcine reproductive and respiratory syndrome virus (PRRSV). The GP5 protein has been known to be a 24.5-26kDa N-glycosylated envelope protein. The GP5 is involved in inducing neutralizing antibodies. For this reason, the GP5 is primary candidate for the PRRSV subunit vaccine. To produce the native form of GP5 in mammalian cells, we have cloned the ORF5 gene from PRRSV CNV-1 into the Semliki Forest virus (SFV)-based expression vector, resulting in recombinant pSFV-ORF5. By the infection with recombinant pSFV-ORF5 to BHK-21 cells, the GP5 expression was confirmed by immunocytochemistry and immunoblotting assay. The recombinant virus particle harboring ORF5 gene was infectious to BHK-21 and MARC-145. The RNA synthesis and expression of GP5 in the infected cell was also confirmed by RT-PCR.

Keyword

PRRS virus; ORF5 gene; GP5; Semliki Forest virus-based expression vector; recombinant SFV particle

MeSH Terms

Animals
Base Sequence
DNA Primers
*Genes, Viral
Plasmids/genetics
Porcine respiratory and reproductive syndrome virus/*genetics
Restriction Mapping
Reverse Transcriptase Polymerase Chain Reaction
Semliki forest virus/*genetics
Swine
Viral Envelope Proteins/genetics
Viral Proteins/*genetics
Virology/methods