Mink parvovirus virus-like particle as well as preparation method and application thereof

A mink parvovirus, virus-like technology, applied in the field of veterinary biopharmaceuticals and microorganisms, to achieve the effects of good immunogenicity, good protection and high titer

Active Publication Date: 2015-03-11
CHANGCHUN SR BIOLOGICAL TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, if the four amino acid sequences of polyhedrin protein are fused and expressed in mink parvovirus VP2, can the exogenous protein be expressed at a high level in Spodoptera frugiperda ovary cells (Sf9 cells), and whether the exogenous protein can be correctly packaged into virus-like particles , and the level of immunogenicity and reactogenicity, etc., there is no literature support

Method used

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  • Mink parvovirus virus-like particle as well as preparation method and application thereof
  • Mink parvovirus virus-like particle as well as preparation method and application thereof
  • Mink parvovirus virus-like particle as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Example 1 Mink parvovirus virus-like particles and its preparation method

[0030] 1 material

[0031] Mink parvovirus, strain E.coli DH10Bac, Sf9 cells, and fetal bovine serum were provided by Changchun Sino Biotechnology Co., Ltd., pEASY-Blunt Simple vector was purchased from Beijing Quanshijin Biotechnology Co., Ltd., pFastBac Dual vector, liposome 2000 was purchased from Invitrogen Company, Phusion DNA polymerase, restriction endonuclease and T4 DNA ligase were purchased from NEB Company, IPTG and X-gal were purchased from Dalian Bao Biological Company, antibiotics were purchased from Sigma Company, Grace insect cell culture medium, Sf -900II SFM serum-free cell culture medium was purchased from Gibco Company, cell culture dishes and cell shaker flasks were purchased from Corning Company, and DNA extraction kits, gel recovery kits, and plasmid extraction kits were purchased from Axygen Company.

[0032] 2 methods

[0033] 2.1 Extraction of virus genome

[0034] I...

Embodiment 2

[0051] Example 2 Identification of Mink Parvovirus Virus-Like Particles and Immunization of Mink

[0052] 1 material

[0053] Fresh pig blood, rabbit serum, and parvovirus monoclonal antibody were provided by Changchun Sino Biotechnology Co., Ltd. FITC-labeled goat anti-mouse secondary antibody and HRP-labeled goat anti-mouse IgG were purchased from Sigma Company.

[0054] 2 methods

[0055] 2.1 Identification of Mink Parvovirus VLPs

[0056] 2.1.1 Hemagglutination test

[0057] Using fresh pig blood, the buffer solution is 15mM PBS with pH 6.5 for the determination of hemagglutination titer. The steps are as follows: centrifuge the freshly taken pig blood preserved in Alfred's solution at 5000rpm for 5min, wash it with PBS three times, after the centrifuged supernatant is clarified, prepare 1% erythrocytes from the centrifuged erythrocytes, and add 0.5% rabbit serum. Add 25 μL of PBS to the 96-well V-type hemagglutination plate, add 25 μL of the sample to be tested, and p...

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Abstract

The invention provides a mink parvovirus virus-like particle which is characterized in that VP2 gene of mink parvovirus is optimized according to insect cell optimal codons, the 5' end of the optimized VP2 gene is directly connected with the nucleotide sequence of polyhedron of a coding part and then is cloned into a transfer vector, and the mink parvovirus virus-like particle can be produced through a baculovirus / insect cell expression system. The mink parvovirus virus-like particle can be produced through the expression system in a safe, efficient and large-scale manner, and the obtained virus-like particle is high in titer and good in immunogenicity; both muscle immunization and oral immunization can induce a mink body to produce high-level specific antibody which can resist attack of virulent virus and well protect a mink, and a foundation is laid for the preparation of mink viral enteritis vaccine.

Description

Technical field [0001] The present invention relates to the fields of microorganisms, genetic engineering and veterinary biopharmaceuticals, and specifically to mink parvovirus virus-like particles and their preparation methods and applications. Background technique [0002] Mink viral enteritis is a severe infectious disease caused by mink enteritis viurs (MEV), also known as infectious enteritis or panleukopenia, in cats, mustelids, minks and canines. All are susceptible, and it is one of the three major viral infectious diseases that poses a greater threat to the mink breeding industry. The disease was first discovered by Schofield in a mink farm in Fort William, Canada in 1949. In 1952, Will extracted the pathogen and named it MEV. Subsequently, the disease quickly spread to the United States, Denmark, Norway, Sweden, the Soviet Union, Japan and many other countries. The disease was first discovered in my country in 1974 and has since spread across the country, causing...

Claims

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Application Information

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IPC IPC(8): C07K19/00C12N15/62C12N15/866C12N7/04A61K39/23A61P31/20A61P1/00C07K16/08C12R1/93
Inventor 金宏丽夏晓红陈宪平付玉刘冰杨佳夏振强石晶
Owner CHANGCHUN SR BIOLOGICAL TECH
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