Eukaryotic Expression Of Cpgp40/15 And Its Immunoprotective Study

The Cpgp40/15 on the sporozoites of Cryptoporidiwn parvum are major surface protein .It is localized to the surface and apical region of invasive stages of the parasite and is shed from its surface.The surface localization of Cpgp40/15 and its involvement in the host-parasite interaction suggest that the glycoprotein may serve as effective targets for specific preventive or therapeutic measures for cryptosporidium.In the experiment, eukaryotic expression vectors of Gp40 genes was constructed by recombinant DNA technique and expressed in Hela cells respectively.The nucleic acid vaccine could induced protective immune responses against C.parvum after BALB/c mice was inoculated intranasally.Cloning and Nucleotide Sequencing of Cpgp40/15 Cpgp40 gene was amplificated from genomic DNA of Cryptosporidium parvum by polymerase chain reaction (PCR), .PCR product was inserted into vector pMD18-T direct!vely. Comparison and analysis of the nucleotide and deduced amino sequences was performed with that reported in Genebank, Then they were sequenced by Sanger's method. The results showed the homology of the nucleotide and amino acid sequences were 98% and 98%.Construction and expression of eukaryotic expression vectors The recombinant plasmids pVAXl-Cpgp40/15 was contructed by cloning Cpgp40/15 genes into eukaryotic expression vector pVAX1 and expressed in Hela cell strain.After transfected it into Hela cells, expression cell strain were selected with G418.The specific recombiant proteins were detected by SDS-PAGE and Westen-blot methods.The protective immune response induced by ncleic acid vaccine in mice All the BALB/c mice were immunized with the vaccine plasmids by different methods The responses of specific systemic and mucosal immunity were elevated by the immunological methods such as the ratio of CD4+/CD8+ and the specific antibodies responses were strengthened with the increase of immunization times.The immunity indexes between experiment groups were not significant,while those between experiment and control groups were significant.The results also indicated that immunized with nucleic acid vaccine was very good.l...