Construction of a gE/gI-deleted mutant strain of epidemic porcine pseudorabies virus and its biological characteristics

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.