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Genetic Stability and Immunizing Dose of Recombinan Fowl Poxvirus Expressing E0-E2 Gene of Classical Swine Fever Virus

Author: ZhangHao
Tutor: ZhangYanMing
School: Northwest University of Science and Technology
Course: Preventive Veterinary Medicine
Keywords: Swine fever Recombinant fowlpox virus Genetic stability Immunization dose
CLC: S852.65
Type: Master's thesis
Year: 2008
Downloads: 69
Quote: 1
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Abstract


Swine fever (Classical swine fever, CSF) is a pig and wild boar caused by the classical swine fever virus (CSFV) a highly lethal and contagious disease, is a serious threat to the global epidemic, one of the pig industry. Immunization of swine fever vaccine is an effective means of prevention of swine fever. Classical swine fever vaccine candidate of CSFV E0-E2 recombinant fowlpox virus live vector vaccine virus seed genetic stability and vaccine dose study, and obtained the following results. 1 recombinant fowlpox virus rFPV-E0-E2 on chicken embryo fibroblasts continuous passage for 20 generations, take 5, 10, 15 and 20 generation of recombinant viruses, respectively, with a blue plaque assay, PCR amplification and gene sequence analysis and indirect immunofluorescence test the genetic stability of the recombinant virus. Results of generations of recombinant virus plaques are blue, from each generation of recombinant viral DNA was amplified E0 and E2 gene, gene sequence analysis revealed two point mutations in only 20 generations recombinant virus inserted gene E0 gene 139A → G, E2 gene 413A → G. The indirect immunofluorescent also be able to detect the expression of the target protein. The results showed that the recombinant fowlpox virus rFPV-E0-E2 in chick embryo fibroblasts uploaded within 20 generations with good genetic stability of CSFV E0-E2 gene insertion to correct stable expression. CEF cells amplified three batches of recombinant fowlpox virus rFPV-E0-E2, and its purity, the virus content, the expression of the specific protein detection and sterility testing. The results showed that three batches of recombinant virus rFPV-E0-E2 has a higher purity, free of bacterial and fungal contamination, the TCID 50 of 10 -7.5 / 0.025 mL, 10 -9 / 0.025 mL and 10 -9 / 0.025 mL, and were able to properly express the of CSFV E0-E2 gene intended as a vaccine original drug all the sub-batches. Meanwhile, laboratory production and the inspection of the batch of swine fever virus E0-E2 recombinant fowlpox virus live vector vaccine virus, the measured LD CSFVShimen strains of the 50 to 60-day-old swine fever susceptible pigs 50 10 -2.8, / 2mL swine fever virus E0-E2 recombinant fowlpox virus immunization dose trials material basis. 3. Use the variable immune quantitative challenge method CSFV recombinant fowlpox virus immunization dose trial. 23 pigs were randomly divided into six groups, each of the different procedures Immune different dose of recombinant fowlpox virus. After immunization 28d, each pig neck intramuscular injection of 100 LD 50 Shimen strain of CSFV continue to observe 21 days to evaluate the protective effect of vaccine. By the 100LD 50 CSFVShimen strains intramuscular injection attacks results to 1.04 × 10 7 pfu a of CSFV recombinant fowlpox virus rFPV-E0-E2 vaccinated twice pig can 50% of the clinical protection that the vaccine CSFV challenge clinical symptoms of PD 50 2 × 1.04 × 10 7 pfu. The results provide a basis for determining vaccine dose.

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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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