Fundamental Study On Recombinant Living Vectored Vaccine Based On Canine Adenovirus Type 2 With Taenia Solium Oncosphere TSOL18 Gene

Cysticercosis is an important parasitic zoonosis ,caused by the larval stage of Taenia Solium, Cysticercus cellulosae in pigs or human beings.Human is not only a intermediate host but also a terminal host.Adenovirus is an important gene transfer vector in research of engineering vaccines, development of TSOL18 DNA vaccine and living vectored vaccine based on Canine Adenovirus type 2(CAV-2) will be great value in control or eradication of cysticercosis in the world.In order to evaluate the protection of DNA vaccine and living vectored vaccine based on CAV-2of TSOL18 gene from Taenia solium oncosphere,we started fundamentally this study.According to Kozak rule, a pair of primers used to amplification of TSOL18 gene from the recombinant plasmid pGEM-TSOL18 by PCR was designed and synthesized .The PCR products digested by EcoR I and Xho I were ligated into pVAX1 and the positive was named pV-TSOL18.Moreover,two other pairs of primers were designed according to the method of PCR site mutation.After three PCRs, mutated TSOL18 gene(TSOL18m) was inserted into the downstream of CMV promoter of pVAXl to obtain the recombinant vector pV-TS0L18m providing the basis of development of recombinant living vectored vaccine.The transcriped TS0L18m gene in BHK-21 Cells was measured 36 hours after transfection.In order to construct a recombinant canine adenovirus type 2(CAV-2) to express the TSOL18m protein of Taenia solium oncosphere , total expression cassette including pCMV,TS0L18m and BGH polyA was cloned into the shuttle vector pVAXAE3,then further cloned into the backbone vector pPoly2-CAV2 containing complete genome of CAV-2.To obtain the recombinant canine adenovirus ,the recombinant plasmid pCAV2-TSOL18m was linerized by Asc I and Cla I to release recombinant genome,and then transfected into DK cells.The recombinant virus CAV2-TSOL18m was obtained and the classical cytopathic effects(CPE) of CAV-2 could be observed . The transcriped TS0L18m gene was measured by RT-PCR analysis.