Abstract:
Objective To obtain a candidate vaccine strain against epidemic porcine pseudorabies virus.
Method A gE/gI-deleted transferring plasmid pMD-LA-RA and a recombinant plasmid carrying EGFP gene were constructed according to the sequence of epidemic porcine pseudorabies virus. The homologous recombination was operated between pMD-LA-EGFP-RA and PRV AH, and then the recombinant mutant virus PRV AH gE –/gI –/EGFP+ was selected using EGFP as screening marker. In order to obtain the gE/gI-deleted mutant strain PRV AH gE –/gI – without the EGFP gene, the second homologous recombination was carried out between pMD-LA-RA and PRV AH gE –/gI –/EGFP+. The proliferation ability, genetic stability and immunogenicity of PRV AH gE –/gI – were evaluated by its growth curve, continuous passage in susceptible cells and animal immunization.
Result PRV AH gE –/gI – was obtained through two homologous recombinations with fluorescence observation, plaque purification and PCR. The deletion of gE, gI genes and EGFP gene marker in PRV AH gE –/gI – was identified by PCR, fluorescence observation and sequencing. PRV AH gE –/gI – had similar proliferation ability to the parental strain and had good genetic stability and immunogenicity.
Conclusion A gE/gI-deleted mutant strain of epidemic porcine pseudorabies virus with good immunogenicity was constructed successfully, which provides a basis for the development of gene deleted vaccine targeting epidemic strains.
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