Construction and immunogenicity evaluation of BVDV recombinant adenovirus based on E2 protein in mice
To construct the Ad5-BVDV-E2 using the AdMax adenovirus packaging system and to e-valuate its immune efficacy in mice through intramuscular,intranasal,and oral immunization routes.The full length E2 gene sequence of BVDV-1 SWU-Z6 strain was optimized and synthesized based on the codon bias of HEK293 cells.Ad5-BVDV-E2 was prepared using the AdMax adenovi-rus packaging system.The expression of E2 protein in HEK293 was validated by PCR,the indirect immunofluorescence(IFA)and Western blot.Mice were immunized through intramuscular,in-tranasal,and oral routes,as well as BVDV commercial inactivated vaccine group and PBS blank were used as control groups.The recombinant plasmid pDC316-E2 was constructed by inserting the optimized E2 gene and verified through restriction endonuclease analysis and sequencing.Recombinant adenovirus Ad5-BVDV-E2 was obtained through co-transfection of pDC316-E2 and the adenovirus skeleton plasmid into HEK293 cells.The successful insertion of the E2 gene into the adenovirus vector genome was confirmed through PCR.E2 protein could be correctly expressed by the recom-binant adenovirus confirmed by IFA and Western blot analysis,exhibiting a good biological activity in vitro.In addition,high antibody levels were produced through intramuscular injection,in-tranasal,and oral administration.The antibody titer of the Ad5-BVDV-E2 group reached up to 1∶120 400 after one week of enhanced immunization.The successful preparation of Ad5-BVDV-E2 in-duced efficient humoral immune responses in mice,indicating its potential for clinical application.
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