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Canine parvovirus-like particles and preparation method and application thereof

A canine parvovirus and virus-like technology, applied in the field of microorganisms and veterinary biopharmaceuticals, can solve the problems of virulence reversion and recombination, failure to cause cellular immune response, weak immunogenicity of inactivated vaccines, etc., to achieve good immunity and prevention Efficiency, ease of large-scale production, and high immunogenicity

Active Publication Date: 2013-11-13
CHANGCHUN SR BIOLOGICAL TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, the vaccines used for canine parvovirus are mainly attenuated vaccines and inactivated vaccines, but each has its own shortcomings. For example, the attenuated vaccine has the risk of virulence reversion and recombination, and the inactivated vaccine has weak immunogenicity and cannot cause cellular immunity. Reply, etc.

Method used

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  • Canine parvovirus-like particles and preparation method and application thereof
  • Canine parvovirus-like particles and preparation method and application thereof
  • Canine parvovirus-like particles and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Example 1 Preparation and identification of canine parvovirus virus-like particles

[0025] 1 material

[0026] The donor plasmid pFastBac1 was purchased from Invitrogen, and the strain E.coliDH10Bac, insect cell Sf9, and canine parvovirus were provided by Changchun Sino Biotechnology Co., Ltd.

[0027] pEASY-bluntsimple, IPTG, and X-gal were purchased from Dalian Bao Biological Company, Phusion DNA polymerase, T4DNA ligase, and restriction enzyme were purchased from NEB Company, competent cells were purchased from Takata Company, genomic DNA extraction kit, gel Recovery kits and plasmid extraction kits were purchased from Axygen, liposome Lipofectin2000 was purchased from Invitrogen, ampicillin, tetracycline, kanamycin, and gentamicin were purchased from Sigma, and fetal bovine serum was Changchun West Produced by Nuo Biotechnology Co., Ltd., TC-100 insect cell culture medium was purchased from Applichem Company, rabbit anti-canine parvovirus polyclonal antibody was p...

Embodiment 2

[0061] Preparation of Example 2 Canine Parvovirus Enteritis Vaccine and Animal Immunization Test

[0062] 1 material

[0063] After the aluminum phosphate was prepared to an appropriate concentration with isotonic saline, it was ultrasonically crushed for 20 minutes to prepare a suspension, which was sterilized at high temperature for later use. Female guinea pigs (about 250g), Chinese pastoral dogs (2-4 months old, CPV hemagglutination inhibitory antibody titer less than 1:2 3 ) were purchased from Changchun Institute of Biological Products.

[0064] 2 methods

[0065] 2.1 Preparation of canine parvovirus enteritis vaccine

[0066] Inoculate Sf9 cells with recombinant baculovirus expressing canine parvovirus VP2 gene, and culture continuously at 27°C for 4-5 days. After the cells become lesions, harvest the cell suspension, freeze and thaw three times, centrifuge at 2000rpm for 10min, and harvest the supernatant , to obtain canine parvovirus virus-like particles. Canine ...

Embodiment 3

[0082] Example 3 The protective test of canine parvovirus enteritis vaccine to dogs

[0083] 1 material

[0084] Canine parvovirus enteritis vaccine was prepared according to the method described in Example 2, and the Chinese pastoral dog (2-4 months old, CPV hemagglutination inhibitory antibody titer was less than 1:2 3 ) were purchased from Changchun Institute of Biological Products.

[0085] 2 methods

[0086] 2.1 Animal immunity

[0087] Take 12 Chinese pastoral dogs, which are labeled M1, O2, O3, and C4 in turn, and grouped and immunized according to Table 3 in Example 2. Among them, the M1 group received intramuscular injection, and the O2 and O3 groups received oral immunization, with the first immunization on the 0th day and the booster immunization on the 15th day.

[0088] 2.2 Animal infection

[0089] Canine parvovirus virulence challenge was performed 21 days after immunization. The specific method is as follows: oral inoculation of canine parvovirus virulenc...

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Abstract

The invention provides canine parvovirus-like particles. The VP2 gene of canine parvovirus is optimized according to the preferred codons of insect cells, the optimized VP2 gene is connected with an insect expression vector, and the canine parvovirus-like particles are produced by use of a rhabdovirus / insect cell expression system. The canine parvovirus-like particles are combined with a preservative and an adjuvant to prepare a canine parvovirus enteritis vaccine. By adopting the canine parvovirus-like particles provided by the invention, the prepared canine parvovirus enteritis vaccine has the advantages of high output, low production cost, high immunogenicity, good safety, convenience in large-scale production and the like, and has good immunization and prevention effects on canine parvovirus enteritis.

Description

technical field [0001] The invention relates to the field of microorganisms, genetic engineering and veterinary biopharmaceuticals, in particular to canine parvovirus virus-like particles, its preparation method and application. Background technique [0002] In 1978, Australian Kelly and Canadian Thomson isolated the pathogen of canine parvovirus from dogs suffering from hemorrhagic enteritis. In order to distinguish it from canine parvovirus, the isolated pathogen was named CPV-2. In 1979 and 1984 , and two subtypes of canine parvovirus, CPV-2a and CPV-2b, appeared one after another and replaced CPV-2, which are widely prevalent in the world. Reports on CPV-2c appeared in 2002. CPV infects canines and is highly contagious, mainly causing hemorrhagic diarrhea and myocarditis, with high morbidity and mortality in puppies. The disease has become an important infectious disease of canines, causing great harm to the economic animal breeding industry. [0003] Canine parvoviru...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/35C12N15/866C12N7/04A61K39/23A61P31/20
Inventor 夏振强金宏丽郑文文张渭蛟
Owner CHANGCHUN SR BIOLOGICAL TECH
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