Construction Of FMD Bivalence Recombination Fowl Pox Virus And The Immune Study Of Mouse

Foot-and-mouth disease(FMD) caused by FMD virus(FMDV) is one of the most contagious animal virus diseases.At present,FMD is spread in all the world.The virus exists in the form of seven different serotype: O,A,C,Asial,and South African Territorsl(SAT1),SAT2 and SAT3. Serotype O,A and Asial FMD once happened in history of China,and are often epidem c in the country around China and Southeast Asi.Our country has been threatened by Serotype O and Asial FMD at present.Now be used for vaccine advanced recombinant virus vector contain poliovirus,tobacco mosaic virus,baculovirus,fowl pox virus.Fowl pox virus transfer vector to possess other vector unexampled dominance to embody:fowl pox virus genome structure huge and insurmountable,to contain comparatively large exogenous gene;foreign protein may be exactly expressed and in infection cell to gain piously modification.As if glycosylation,carboxylation etc;exogenous gene expression product to possess satisfactory immunogenicity,can induction organism produce last time cytoimmunity and humoral immunity;strict intracytoplasm endoradiosonde,to avoid virogene recombination to enter host cell chromosomal probability,eliminate recombinant virus to be applicated for human and livestock potential threat.The research institute use fowl pox virustransfer vector is artificial composite in-line vaccinia virus mutant p7.5 promoter cowpox virus A type elementary body(ATI) promoter constitutive composition promoter as priming exogenous gene promoter,it may consumedly raise exogenous gene expression quantity.The researcher from contain FMDV cattle blister liquid get out celI RNA,apply RT-PCR amplification Asia-I/JL strain FMDV capsid protein precursor and proteolytic enzyme 3C gene,and to take it cloning pMD18-T vector,through to recombinant plasmid enzyme cutting appraisement,successfully clone FMDV capsid protein precursor and proteolytic enzyme 3C gene,its sides are 2223bp and 639bp,and construction DNA recombinant plasmid T-P1-2A-3C,through enzyme cutting to obtain P1-2A-3C gene expression box.Base on the Military Veterinary Institute doctor Zhang construction O type FMDV clone Plasmid T-P1-2A, Plasmid pUMT-IL18,construction DNA recombinant plasmid T-P1-2A-IL18, through enzyme cutting to obtain P1-2A-3C gene expression box.To take enzyme cutting to gain FMDV Asia-I type P1-2A-3C and FMDV O type P1-2A-IL18 are inserted into the fowl pox virus(FPV) expression vector pUTA-16-LacZ to produce recombinant expression plasmid pUTAL-P1-2A-IL18-P1-2A-3C,The recombinant expression plasmid are then co-transected into CEF cells.The recombinant fowl pox viruses-P1-2A-IL18-P1-2A-3C(rFPV-P1-2A-IL18-P1-2A-3C) is produced by three Cycles with the BrdU and verified by RT-PCR and IFA Western-blot. BALB/c mice were immunized with the recombinant virus by muscular inoculation,and the level of antibodies,the number of spleen T lymphocyte subgroups and cytotoxicity activity of specific CTL were examined after three times every week.The results shoved that both of the recombinant fowl pox viru could increase the percentage of T cell subgroups and elicited cytotoxicity activity of specific CTL which were significantly higher than that of the control.The immunized mice could produce the antibodies against serotypes O and Asia-I FMDV.