Construction And Immunogenicity Of A Retrovirus Containing The Glycoprotein Gene Of Rabies Virus

Recombinant retroviral vector constructed on normal retroviral vector, a kind of replication-defective viral vector, can be significantly multiplied in package cell lines. Exogenous gene is integrated into host cell after infection for its continuous, stable expression. Moreover, the immunized animal may obtain constant immunoprotection.In this research, the rabies virus'protective antigen gene was cloned inside the two long terminal repeat﹙LTR﹚of the retroviral vector pLNCX. Recombinant viruses which were subsequently used to immunize subject animal were harvest from the package cell lines after transfection. The expression of the target gene and the specific antibody level and its rise and fall of the animal subject were determined by multiple methods.The glycoprotein gene of SRV9 strain gained via RT-PCR was cloned into pMD18-T vector for sequencing to determine its correctness. The target gene was then inserted into eukaryotic expression vector pVAX1 to construct integral glycoprotein expression box which was subsequently inserted inside the two LTRs of the retroviral vector pLNCL. After a deletion of promoter CMV, the recombinant plasmids pLNC-G were transfected into package cell line PA317 induced with liposome. Stably expressed cell lines were gained after screening with G418 at the concentration 400μg/mL. Recombinant viruses were harvest from cell lines with higher titers for the animals'immunization. Sera were collected once per two weeks from mice which were immunized by intramuscular injection or intraperitoneal injection. The titers of antilyssic neutralizing antibody in the sera were determined by FAVN.The integration and constant passage of the glycoprotein expression gene were improved by PCR, and the constant transcription was determined by RT-PCR. Animal subjects could produce satisfactory immune response after recombinant virus'induction. Both intramuscular injection and intraperitoneal injection could induce the production of antilyssic neutralizing antibodies. The seroconversion rates were respectively 6.7% and 86.7%.