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O-type foot-and-mouth disease multi-epitope vaccine with cross immunity protective efficiency

A foot-and-mouth disease, multi-epitope technology, applied in the field of biotechnology genetic engineering, can solve the problems of weak immunogenicity, low protective effect, difficult preservation, etc., and achieve the effect of preventing infection

Active Publication Date: 2012-03-14
QINGDAO MINGQIN BIOLOGICAL TECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, these methods also have the problems of weak immunogenicity and low protective effect
The known adenovirus vaccine has a good protective effect, but due to safety problems and storage difficulties in this method, people have not been able to accept it

Method used

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  • O-type foot-and-mouth disease multi-epitope vaccine with cross immunity protective efficiency
  • O-type foot-and-mouth disease multi-epitope vaccine with cross immunity protective efficiency
  • O-type foot-and-mouth disease multi-epitope vaccine with cross immunity protective efficiency

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1 Design ideas for polypeptide-encoded protein of O-type foot-and-mouth disease multi-epitope vaccine

[0043] Integrating the genome sequences, antigen structure and epidemiological research progress of OZK93, OR80, OS99, OHK93 and other strains of O-type foot-and-mouth disease virus epidemic strains in China, the recombinant porcine O-type foot-and-mouth disease multi-epitope vaccine was optimally designed. The present invention uses DNAStar and ANTHEPROT software to analyze the hydrophilicity, antigenicity, plasticity, surface accessibility and Garnier-Robson secondary structure of its main outer membrane proteins VP1, VP2, VP3, and predict possible B cells On the basis of antigenic epitopes and killer T cell epitopes, according to the similarity of epitope positions and amino acid sequences, analyze the common and specific antigenic epitopes of various strains, and refer to the sequence information in GenBank to predict the The antigenic epitopes are compare...

Embodiment 2

[0045] Embodiment two Escherichia coli expression vector and the construction of expression bacterial strain

[0046] Send the designed polypeptide encoding nucleotides in Example 1 to Shanghai Handsome Biotechnology Co., Ltd. for synthesis, and design EcoRI (5' end) and HindIII (3' end) restriction enzyme sites at both ends of the fragment, and then The synthetic fragment was cloned into the pMD18T vector. Sequence determination confirmed that the inserted gene fragment was consistent with the designed sequence (see sequence listing). Using restriction endonucleases EcoRI and HindIII, the O-type foot-and-mouth disease vaccine gene was excised from the vector, and recovered by electrophoresis. The Escherichia coli expression vector was pRSETB plasmid from pharmacia biotech company, also treated with restriction endonucleases EcoRI and HindIII, enzyme digestion conditions: 10 μl reaction system, 2 μl plasmid was added in the system, and the restriction endonuclease was 5 active ...

Embodiment 3

[0051] Example 3 Fermentation, purification and emulsification of engineering bacteria

[0052] Fermentation Take the production strains, inoculate them in 2ml LB liquid medium (containing 100μg / ml ampicillin), and culture at 37°C with shaking at 180rpm for 12 hours to activate the strains. Then inoculate the shake flask with an inoculation amount of 1:100, shake and culture at 37°C until OD600=3, and then inoculate it into a fermenter at a ratio of 10%. The medium used for fermentation is a semi-synthetic medium prepared with distilled water and does not contain any antibiotics. Calibrate the dissolved oxygen and pH electrodes, start the tank to stir, the rotation speed is 300rpm, and sterilize the tank on-line. When the temperature of the culture solution in the tank drops to 37.0°C, calibrate the pH and dissolved oxygen (OD) zero point. The fermentation temperature is 37.0±0.1°C, the dissolved oxygen is controlled at about 20%, and the pH is controlled at 7.0. When the OD6...

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Abstract

The invention relates to preparation and application of a multi-epitope vaccine with cross immunity protective efficiency to O-type foot-and-mouth disease viruses such as OZK / 93, OR / 80MF8, OS / 99MF8, OHK93 and the like. The vaccine contains two sections of T cell auxiliary antigenic epitope polypeptides, 4-7 sections of antigenic epitope polypeptides related to main outer membrane proteins VP1, VP2 and VP3 of different foot-and-mount disease strains. The invention also relates to a preparation method of the vaccine and a clinical immunology application method. The vaccine has stable production preparation process and is applicable to mass production, experiments show that the multi-epitope vaccine is safe to use, infection of different O-type foot-and-mouth circulating strains can be effectively prevented and effective antibody titer can last for at least seven months.

Description

technical field [0001] The invention belongs to the field of genetic engineering of biotechnology, and mainly relates to the preparation and application of an O-type foot-and-mouth disease multi-epitope vaccine with cross-immunity protection effect. Specifically, using gene recombination technology, multiple B cell antigen epitopes, killer T cell antigen epitopes and multiple T cell helper antigen epitopes of the main outer membrane proteins VP1, VP2, and VP3 of different O-type foot-and-mouth disease epidemic strains The method is connected in series, connected to a carrier, transformed into a host bacterium, and prepared through fermentation, purification and emulsification processes to obtain an O-type foot-and-mouth disease multi-epitope vaccine and the application of the vaccine in preventing foot-and-mouth disease. Background technique [0002] Foot-and-mouth disease (FMD) is an acute, febrile, highly contagious infectious disease of artiodactyls caused by Foot and Mou...

Claims

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Application Information

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IPC IPC(8): C07K14/09C07K7/06C07K7/08C07K19/00C12N15/42C12N15/63C12N1/15C12N1/19C12N1/21C12N5/10A61K39/135A61K38/16A61P31/14
Inventor 李殿明蒲勤李毅田春辉赵明齐春梅顾富香任百亮张导春牛纪涛刘甜甜刘祯
Owner QINGDAO MINGQIN BIOLOGICAL TECH CO LTD
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