Bovine A-type foot-and-mouth disease multi-epitope vaccine, and preparation method and application thereof

A foot-and-mouth disease vaccine and multi-epitope technology, applied in the field of veterinary vaccine research, can solve the problems of single antigen spectrum and low immune efficacy of epitope vaccines, and achieve the effect of protection against attack and good application prospects

Inactive Publication Date: 2014-10-29
LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] The present invention adopts molecular biology technology and advanced design concept, aims at the main antigenic epitope sequence of my country's bovine type A foot-and-mouth disease virus strain, develops bovine type A foot-and-mouth disease broad-spectrum multi-epit

Method used

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  • Bovine A-type foot-and-mouth disease multi-epitope vaccine, and preparation method and application thereof
  • Bovine A-type foot-and-mouth disease multi-epitope vaccine, and preparation method and application thereof
  • Bovine A-type foot-and-mouth disease multi-epitope vaccine, and preparation method and application thereof

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Experimental program
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Embodiment 1

[0028] The preparation of embodiment 1 bovine type A foot-and-mouth disease multi-epitope recombinant antigen

[0029] 1. Bioinformatics analysis of VP1 gene of type A foot-and-mouth disease virus:

[0030] The structural protein VP1 of foot-and-mouth disease virus is the dominant antigen of the virus. Whether it is the isolated and purified natural VP1 protein or the recombinant expression product, it can induce the body to produce protective neutralizing antibodies, which is type-specific. The VP1 gene of type A foot-and-mouth disease virus has a full length of 639 nucleotides, encoding a protein of 213 amino acids, and its dominant antigenic epitope is 140-160 amino acids and 200-213 amino acids. In this study, DNAStar biological software was used to analyze the sequences of representative strains A / WH / 09, XJKT58 and GS / LX / 62 of bovine type A foot-and-mouth disease virus recently isolated in my country, and the dominant antigenic epitope was determined to be amino acids 135...

Embodiment 2

[0036] Example 2 Preparation of Bovine Type A Foot-and-Mouth Multi-epitope Vaccine and Analysis of Immune Efficacy

[0037] 1. Vaccine preparation:

[0038] The purified protein (shown in SEQ ID NO.6) was quantified by the Bio-Rad quantitative kit and diluted to an appropriate concentration, alone or with the purified 3D protein full-length fragment (the amino acid sequence shown in SEQ ID NO.8, encoding its The nucleotide sequence is shown in SEQ ID NO.7), and the recombinant antigen was configured at 1:2 (V / V), and an equal volume of oil adjuvant ISA206 (France) was added to emulsify into a vaccine preparation.

[0039] 2. Guinea pig immune efficacy test:

[0040] Using the prepared vaccine, 0.2ml of each guinea pig was inoculated by intramuscular route (20μg antigen or 20μg antigen + 10μg 3D protein) to immunize 5 guinea pigs respectively. The results of the immunization experiment showed that no matter whether the recombinant antigen alone or combined with 3D protein immu...

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Abstract

The invention discloses a bovine A-type foot-and-mouth disease multi-epitope recombinant vaccine, and a preparation method and an application thereof, and belongs to the field of veterinary vaccine research. The preparation method comprises the following steps: carrying out reasonable serial connection on the dominant antigen epitopes of bovine A-type foot-and-mouth disease virus representative strains once pandemic in China by adopting an antigenized antibody strategy and a brand new reverse vaccinology technology and strategy, coupling with a bovine IgG immunostimulatory fragment (IgG heavy chain constant region), cloning into a prokaryotic expression vector to construct a recombinant expression vector, transforming Escherichia coli cells to express a recombinant antigen, purifying by adopting Ni-NAT column chromatography, quantifying by a Bio-Rad protein quantification kit, and preparing the vaccine individually or combining with a recombinant foot-and-mouth disease virus 3D protein. A result of animal immune experiments shows that the recombinant protein or combined vaccine can stimulate the body to produce a protective antibody with high titer and also can protect immune animals against a virus attack, so the bovine A-type foot-and-mouth disease multi-epitope recombinant vaccine has a good application prospect.

Description

technical field [0001] The invention relates to a bovine type A foot-and-mouth disease vaccine and its preparation method and application, in particular to a bovine type A foot-and-mouth disease multi-epitope recombinant vaccine and its preparation method and application, belonging to the field of veterinary vaccine research. Background technique [0002] Foot-and-mouth disease is a major animal disease of pigs, cattle and sheep as the main economic livestock, which 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 political impact will be bad. Inactivated vaccines play a very important role in the prevention and control of FMD outbreaks. However, the production of such vaccines requires the construction of high-level biosafety production workshops to prevent pathogens from escaping, and to distinguish between vac...

Claims

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Application Information

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IPC IPC(8): C07K19/00C12N15/62C12N15/63C12N5/10A61K39/135A61K48/00A61P31/14
Inventor 邵军军常惠芸陈建文
Owner LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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