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Porcine foot-and-mouth disease virus O-type fc polypeptide vaccine and its preparation method and application

A technology for swine foot-and-mouth disease virus and polypeptide vaccine, which is applied in the field of swine foot-and-mouth disease virus O-type Fc polypeptide vaccine and preparation thereof, and the field of swine foot-and-mouth disease vaccine and its preparation, can solve the problems of high cost and complicated preparation process, and achieves high cost and immunogenicity. Strong, low recovery effect

Active Publication Date: 2021-07-27
LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The technical problem to be solved by the present invention is to overcome the problems such as complex preparation process and high cost of the existing porcine foot-and-mouth disease virus O-type broad-spectrum multi-epitope vaccine, and provide a kind of porcine foot-and-mouth disease virus O-type Fc polypeptide vaccine after optimization and improvement thereof. Preparation

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Codon optimization of embodiment 1 porcine foot-and-mouth disease virus O type broad-spectrum multi-epitope fusion gene

[0030]Using bioinformatics code optimization software combined with multidisciplinary technologies such as immunology and biochemistry, the patent application with the publication number CN102675471A and the invention title "Porcine Foot-and-Mouth Disease Virus Type O Broad-spectrum Multi-epitope Recombinant Antigen and Its Application" was disclosed The nucleotide sequence encoding the porcine foot-and-mouth disease virus type O broad-spectrum multi-epitope recombinant antigen is codon-optimized, and the optimized nucleotide sequence is shown in SEQ ID NO.1. The sequence comparison analysis (Clustal W software and DNAMEN Version 9) of the genes before and after optimization showed that there were large mutations in the gene sequence before and after optimization in the 1-684bp region, and there were basically no major mutations in the Fc segment of I...

Embodiment 2

[0031] Example 2 Cloning of the gene encoding porcine foot-and-mouth disease virus O-type Fc polypeptide and its protein expression, purification

[0032] In order to ensure directional insertion, specific enzyme cutting sites such as BamHI and XhoI were introduced at the 5'-end and 3'-end of the fusion gene shown in SEQ ID NO.1, and entrusted to Suzhou Jinweizhi Biotechnology Co., Ltd. to synthesize. The synthetic gene encoding porcine foot-and-mouth disease virus O-type Fc polypeptide was inserted into the prokaryotic expression vector pET-28a(+) to construct the recombinant expression plasmid pMEO-Fc, transformed into JM109 competent for positive screening, and confirmed by BamHI and XhoI double enzyme digestion and sequence determination Positive recombinants were stored at -20°C for future use.

[0033] Expression of recombinant protein and identification of its biological activity: Transform the positive recombinant expression plasmid pMEO-Fc into BL21(DE3)pLysS(Novagen)...

Embodiment 3

[0034] Example 3 Vaccine preparation and immune efficacy test

[0035] 1. Vaccine preparation

[0036] Vaccine of the present invention: the purified protein obtained in Example 2 is diluted to an appropriate concentration after being quantified by the Bio-Rad quantitative kit, and is emulsified by adding oil adjuvant Montanide ISA206 (Seppic, France) according to a 1:1 (w / w) ratio. Vaccine preparation (W / O / W), 1ml per head, containing 200μg of soluble porcine foot-and-mouth disease virus O-type Fc polypeptide.

[0037] Control vaccine: Purified recombinant antigen and purified full-length 3D protein were prepared according to the method disclosed in CN102675471A. They were quantified by Bio-Rad quantitative kit and then diluted to an appropriate concentration. : After 2 (V / V) preparation, add an equal volume of oil adjuvant Montanide ISA206 (Seppic, France) to emulsify into a vaccine preparation, 1ml per head, which contains 200μg of recombinant antigen and 100μg of full-len...

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Abstract

The invention discloses a porcine foot-and-mouth disease virus O-type Fc polypeptide vaccine as well as a preparation method and application thereof. The porcine foot-and-mouth disease virus O-type Fc polypeptide vaccine contains the soluble porcine foot-and-mouth disease virus O-type Fc polypeptide obtained by expressing the nucleotide sequence shown in SEQ ID NO.1 through a prokaryotic expression system. The present invention optimizes the codon of the disclosed nucleotide sequence encoding porcine foot-and-mouth disease virus O-type broad-spectrum multi-epitope recombinant antigen, and then realizes the solubility of the target protein by rebuilding the recombinant plasmid and optimizing induction and culture conditions. expression and purification. Soluble protein mimics the characteristics of natural antigens to the greatest extent, with stronger immunogenicity and higher antibody levels. In addition, the vaccine prepared by the invention has simpler components, does not contain 3D protein components, reduces the total protein content of the vaccine, and reduces vaccine cost while improving immune efficacy. The proposal of the present invention provides material reserves and technical support for the prevention and control of porcine O-type foot-and-mouth disease in my country, and has important significance.

Description

technical field [0001] The invention relates to a porcine foot-and-mouth disease vaccine and its preparation method and application, in particular to a porcine foot-and-mouth disease virus O-type Fc polypeptide vaccine and its preparation method and application. The invention belongs to the technical field of medicine. Background technique [0002] As a major animal disease, foot-and-mouth disease not only seriously threatens the healthy development of animal husbandry, but also involves the safety of animal-derived food and its foreign trade export. Once an epidemic occurs, the losses will be heavy and the impact will be severe. The World Organization for Animal Health (OIE) lists it as an infectious disease that must be notified, and my country lists it as a first-class animal infectious disease. In order to purify and eliminate foot-and-mouth disease, my country has formulated the "National Medium- and Long-Term Animal Disease Prevention and Control Plan (2012-2020)" and...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/62C07K19/00C12N15/70A61K39/135A61K47/68A61P31/14
CPCA61K39/12A61K2039/552A61K2039/55511C07K14/005C07K2319/30C12N15/62C12N15/70C12N2770/32122C12N2770/32134C12N2800/22
Inventor 邵军军常惠芸李杨帆张永光
Owner LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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