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Generation of Recombinant Baculovirus Expressing VP2 Protein of Mink Enteritis Virus and Analysis of Its Immunogenicity in a Rabbit Model

Author: NiJia
Tutor: ZuoXiJun
School: Chinese Academy of Agricultural Sciences
Course: Preventive Veterinary Medicine
Keywords: Mink enteritis virus Baculovirus expression system Virus-like particles
CLC: S852.65
Type: Master's thesis
Year: 2011
Downloads: 91
Quote: 2
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Abstract


Mink virus enteritis (Mink viral enteritis) is an acute, highly contagious disease, the disease is mainly characterized by diarrhea caused by mink enteritis parvovirus. Its high morbidity and mortality, the world recognized hazards mink husbandry larger viral infectious diseases, supplies mink industry has brought huge economic losses. With other viral diseases, the prevention and treatment of mink viral enteritis also mainly focusing on prevention. The virus-like particle (Virus-like particles) is a highly structured protein particles self-assembled by the the virus single or plurality of structural proteins. In shape, similar to immature virus particles of the virus-like particles, but due to lack of regulatory proteins and the infectious nucleic acid, no replication and infection ability. Despite the lack of infectious virus-like particles with similar natural viral tropism, be the cellular uptake and intracellular transport stimulate overall immune response. In this study, the expression of the baculovirus expression system MEV VP2 gene MEV virus-like particle preparation. In this study, theoretical basis, application baculovirus expression system (Bac-to-Bac system) was constructed to express MEV VP2 protein MEV VP2 protein self-assembled into virus-like particles, and still maintain its immunogenicity The recombinant baculovirus, to establish its basis MEV subunit vaccine preparation method. Mink enteritis virus (MEV) SMPV-11 strain VP2 gene was amplified by PCR, VP2 gene cloning and sequencing analysis showed that the virus attribution of the current the MEV major popular antigen type MEV-1 type. The VP2 Gene SMPV-11 and pFastBac Ⅰ carrier digested cloning connect successfully construct the recombinant plasmid pFastBack-VP2 shuttle vector. The shuttle vector be transformed into DH10Bac, blue-white and resistance screening recombinant VP2 bacmid Bacmid-VP2. The bacmid applications liposomal transfection of Sf9 cells can infect the cells the recombinant baculovirus rBacMEV-VP2. Cells were infected projection electron microscopy confirmed rBacMEV-VP2 in Sf9 cells expressing VP2 protein, protein expression of parvovirus-like particles can be formed. The expression products were confirmed by Western-blotting and indirect immunofluorescence assay VP2 protein expression can be the MEV monoclonal antibody recognition. Recombinant baculovirus expression rBacMEV-VP2 VP2 protein subunit vaccine and purified by centrifugation as aluminum glue the adjuvant preparation of MEV vaccine. The respectively 100μg, 500μg different doses of immune Japanese white rabbits after immunization 2w animal serum-MEV HI antibody positive, 3w antibodies can reach the highest level, respectively 1:108.8,1:204.8. Experiments show that virus-like particles prepared MEV vaccine generated antibody levels below the commercialization of the MEV inactivated vaccine.

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CLC: > Agricultural Sciences > Livestock, animal medicine,hunting,silkworm,bee > Animal Medicine ( Veterinary Medicine) > Basic Veterinary Science > Animal Microbiology ( Veterinary Microbiology, ) > Livestock Virology
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