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Cloning of VP3 Gene of Goose Parvovirus and Construction of Recombinant Fowlpox Virus Transfer Vector

Author: TianLiHong
Tutor: JiaYongQing;WangJunWei
School: Northeast Agricultural University
Course: Preventive Veterinary Medicine
Keywords: Goose Parvovirus VP3 gene Clone Recombinant fowl pox virus transfer vector
CLC: S852.65
Type: Master's thesis
Year: 2002
Downloads: 115
Quote: 1
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Abstract


Goose Parvovirus (Goose Parvovirus GPV) gosling plague pathogens. Gosling plague occurred in the 1-month-old goslings and Muscovy ducklings, based on acute enteritis and liver, kidney, heart parenchymal organs deadly infectious diseases characterized by inflammation, posing a great threat to the development of the goose industry. Since 1956, Chinese scholars side a first discovered and isolated GPV abroad have been isolated reports of the virus, the virus more widely popular in the world. Was amplified by PCR and cloned GPV H1 isolates the main immunogenic protein gene VP3 and prokaryotic and eukaryotic expression is built gosling plague molecular diagnostic methods, to build the basis of VP3 recombinant fowl pox virus live vector vaccine , an extremely important theoretical and practical significance. The virus multiplication methods used in this study were inoculated duck embryo GPV H1 isolates proliferation, proliferation of purified virus by differential centrifugation. Nucleotide sequence according the GPV Write Zadori B strains, one pair of primers for the amplification of a major structural protein VP3 gene GF / GR design means Oligo4.1 software virus genomic DNA was amplified by the PCR technique, from a the virus major structural protein VP3 complete gene fragment, directly connected with the PMD 18-T plasmid vector after restriction enzyme digestion. Extracting the recombinant plasmid are identified by PCR and restriction enzyme digestion, insert sequencing and analysis. The results showed that: the goose parvovirus H1 isolates VP3 gene is 1605bp, encoding 534 amino acids, only a complete open reading frame, published abroad goose parvovirus B strains nucleotide sequence homology of 98.5%, amino acids The homology was 98.3%, indicating that these two strains similar phylogenetic relationship. GPV B strains VP3 gene nucleotide sequence of the sequence published abroad compared to a total of 24 nucleotides. Apart from the individual point mutations, most of the variability is concentrated in two regions, the VP3 gene 5 'end of the 312bp-417bp region and 3' terminal 791bp-1276bp amino acid sequence deduced from the sequence and GPV B strains 9 amino acids change concentrated between 266 and 417 amino acids in this region may VP3 gene hypervariable region. This study also cut from the containing GPV H1 isolates the major structural protein VP3 gene in the recombinant plasmid the pPROEX HTB-VP3 GPV H1 strains VP3 gene fragment subcloned into in pSY538 EcoRI sites, and then with a vaccinia virus promoter P11 SmaI sites of the LacZ reporter gene blunt end cloned in the above-mentioned recombinant, and then the NotI cut containing both VP3 gene and the LacZ reporter gene fragment subcloned ships the pSY681 the NotI sites containing VP3 gene construct recombinant fowlpox virus transfer vector, GPV genetic engineering to prepare for the construction of expression VP3 recombinant fowl pox virus vaccine laid the foundation.

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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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