Construction and identification of a recombinant canine type 2 adenovirus expressing VP2 protein of canine parvovirus

The E3-region-deleted vector, named pVAXDelta E3, was constructed by removing the E3 Ssp Ⅰ segment. The CPV VP2 expression cassette released from pVCPV-VP2 plasmid was ligated into Ssp Ⅰ site of pVAXDelta E3 plasmid, and the resulted recombinant plasmid was named pDelta ECPV-VP2, pAECPV-VP2 and pPoly2-CAV-2 were digested with Nru Ⅰ/Sal Ⅰ, respectively. The purified Nru Ⅰ/SalⅠ DNA fragment containing the CPV VP2 expression cassette was cloned into pPoly2-CAV-2 plasmid to generate pCAV-2/CPV-VP2. The recombinant genome, CAV-2/CPV-VP2, was released from pCAV-2/CPV-VP2 plasmid by digestion with Asc Ⅰ/Cla Ⅰ. CAV-2/CPV-VP2 was cotransfected into DK cells with CAV-2 genome without the Nru Ⅰ/Sal Ⅰ segment by Lipofectamine. The recombinant virus, named CAV-2/CPV, was gained and it produced classical cell cytopathic effects in DK cells. The morphologic character of CAV-2/CPV is similar to CAV-2. Expression of CPV VP2 gene by CAV-2/CPV in infected DK cells was confirmed by RT-PCR. CAV-2/CPV propagated slower than wild CAV-2. Hereditary feature of CAV2/CPV is stable.