Expression and analysis of recombinant VP1 and VP2 capsid protein of swine vesicular disease virus

CH Pan , TH Chen, MK Wang, LC Hung, C Wang, WL Yu, TS Huang

Animal Health Research Institute, Council of Agriculture, Executive Yuan

Abstrate

The purpose of this study was to express the structure proteins of SVDV and to
compare different expression vectors for the production of soluble prokaryotic recombinant proteins.
Different purification systems were used to purify the recombinant proteins and indirect ELISA was
used to test SVD positive pig sera. Full length of VP1 and VP2 genes of SVDV were amplified by
RT-PCR. The RT-PCR amplicons were cloned into a TOPO vector using TA cloning kit; correct
clones were confirmed by sequencing. The target genes were subcloned into the pET28, pET32,
SUMO and pCold expression vectors. The expression plasmids were transformed into host cell of
BL-21 (DE3), then IPTG was used to induce the expression of cloned genes. The solubility of
recombinant proteins was tested after induction. Results showed that only pCold vector expressed a
soluble protein. His tag protein purification and gel extraction by slicing were used to compare the
purity of recombinant proteins. Results demonstrated that gel slicing could obtain a high purity
recombinant protein. The ELISA plates were coated with purified recombinant VP1 protein and
indirect ELISA method was used to test serial dilutions of pig sera which were obtained from the
experimental SVDV infected pigs. Results revealed that linear curve could be observed in the
indirect ELISA assay. A panel of standardized SVD reference pig sera was also tested by indirect
ELISA. Results showed that high positive correlation was observed between ELISA test and virus
neutralization test (VNT).

Keywords: swine vesicular disease (SVD), reverse transcription polymerase chain reaction
(RT-PCR), enzyme-linked immunosorbent assay (ELISA), virus neutralization test (VNT).