Construction And Characteirzation Of A Recombinant Pseudorabies Virus Expressing VP2of Porcine Parvovirus And Cap Of Porcine Circovirus Type2

Pseudorabies virus (PRV), Porcine Parvovirs (PPV) and Porcine circovirus type2arethree important pathogens of swine, resulting in devastating and economic loss worldwide.Pseudorabies virus disease and Porcine Parvovirus disease are infectious swine which inducespregnant swine abortion, stillbirths. Porcine circovirus type2is a swine virus, whichassociated from various disease syndromes in swines and resulting in massive economic lossworldwide.PRV is the best virus as a vector which used to express foreign gene, because of thegenome of PRV DNA about150Kb, which contains many nonessential gene, broadest hostrange.With the gene-deleted pseudorabies virus(PRV) which has been used as the vaccine foreradication Pseudorabies virus disease and a live-viral vaccine vector to develop bivalent ormultivalent genetic engineering vaccine.This study based on the bacterial artificial chromosome of PRV, which the gene of gEhas been deleted, to development of porcine Pseudorabies virus (PRV), Porcine Parvovirus(PPV) and Porcine circovirus type2(PCV2) trivalent inactivated vaccine, the recombinantplasmid pEP-V2C-in was constructed by inserting VP2-2A-Cap (V2C) gene, which is theCDS of porcine parvovirus VP2and porcine circovirus type2Cap connecting by FMDV2Agene, into expression vector pEP-CMV-in. The BAC clones of pPRV-V2C-ΔgE andpPRV-ΔgE were constructed using two Red E/T methods. Then the recombinant virusrPRV-V2C-ΔgE and rPRV-ΔgE were rescued by transfecting the obtained BAC DNAs intoBHK-21cells by calcium phosphate precipitation. Western blotting assay showed that PPVVP2and PCV2Cap proteins were successfully expressed in rPRV-V2C-ΔgE virus-infectedBHK-21cells with the molecular mass of64ku and28ku respectively, which demonstratesthat the fused protein were successfully cleaved by FMDA2A. Furthermore, the results ofgrowth kinetics and plaque formation sizes showed that insertion of the foreign genes had noinfluence on the propagation of rPRV-Bac-EF1in BHK-21cells. These studies have laid afoundation for developing the recombinant PRV vector vaccine expressing PPV VP2andPCV2Cap.