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Recombinant virus secreting multi-epitope gene expressing porcine O-type foot-and-mouth disease virus and preparation method and application thereof

A foot-and-mouth disease and epitope technology, applied in botany equipment and methods, biochemical equipment and methods, medical preparations containing active ingredients, etc., to increase expression, protein secretion, expression and immunogenicity Effect

Active Publication Date: 2019-06-14
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Multi-antigen epitope tandem is a method to improve the immunogenicity of antigenic protein, but there is no relevant report on porcine O-type foot-and-mouth disease virus multi-antigen epitope tandem to prepare vaccines

Method used

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  • Recombinant virus secreting multi-epitope gene expressing porcine O-type foot-and-mouth disease virus and preparation method and application thereof
  • Recombinant virus secreting multi-epitope gene expressing porcine O-type foot-and-mouth disease virus and preparation method and application thereof
  • Recombinant virus secreting multi-epitope gene expressing porcine O-type foot-and-mouth disease virus and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] The construction of the recombinant baculovirus Ac-RBT of embodiment 1 expression multi-epitope recombinant FMDV RBT protein

[0044] (1), acquisition of recombinant baculovirus Ac-RBT

[0045] 1. Bioinformatics analysis of VP1 gene of porcine O-type foot-and-mouth disease virus

[0046] In this study, DNAStar biological software was used to analyze the sequence of the domestic popular strain O / BY / CHA / 2010 (NCBI accession number: JN998085.1), and the dominant B cell antigen epitopes were determined to be 129th to 160th and 200th to The 213th amino acid, the dominant T cell epitope is the 16th to 44th amino acid.

[0047] 2. Design and synthesis of multi-epitope recombinant FMDV RBT gene

[0048] The present invention introduces a spacer sequence GG between adjacent epitopes to ensure that each epitope structure can be displayed correctly. Taking B(129~160)-B(200~213)-B(129~160)-T(16~44) as a basic unit, artificially synthesized 3 basic units of 1122bp in series accor...

Embodiment 2

[0068] Example 2 The immune efficacy experiment of the subunit vaccine prepared by recombinant baculovirus Ac-RBT

[0069] 1. Preparation of recombinant baculovirus Ac-RBT porcine O-type foot-and-mouth disease subunit vaccine

[0070] With the recombinant baculovirus Ac-RBT prepared in Example 1, according to 1M.O.I. infection culture concentration is 2×10 6 cells / mL of Sf9 suspension cells. The culture was collected 72 hours after infection, the culture supernatant was removed by centrifugation, the cells were resuspended by adding an equal amount of PBS buffer, the suspension was processed by an ultrasonic cell disruptor, and cell debris was removed by centrifugation. After measuring the concentration of the target protein in the ultrasonic supernatant, adjust the protein concentration to 100 μg / mL, and then emulsify with ISA201VG adjuvant (SEPPIC company product) to prepare a subunit vaccine. At this time, 50 μg of RBT protein is contained in each ml of vaccine, and stored...

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Abstract

The invention discloses a recombinant virus secreting a multi-epitope gene expressing a porcine O-type foot-and-mouth disease virus, a preparation method thereof and application thereof. The gene sequence of a multi-epitope recombinant antigen of the porcine O-type foot-and-mouth disease fused with bee venom signal peptide is artificially designed according to the preference of a codon by connecting T and B cell epitopes of the porcine O-type foot-and-mouth disease virus, and a recombinant baculovirus containing the above recombinant gene is constructed to increase the secreting expression level and the immunogenicity of the gene in the baculovirus. A subunit vaccine is prepared by using an RBT protein expressed by the recombinant baculovirus Ac-RBT. After a pig is immunized, the body canbe induced to have specific immune reaction, and the pig body is induced to produce a higher level specific antibody which can effectively prevent the foot-and-mouth disease virus, and is of great significance for control and purification of the porcine O-type foot-and-mouth disease in China.

Description

technical field [0001] The invention relates to the technical field of veterinary biological products, and more specifically relates to a recombinant virus secreting and expressing multi-epitope genes of porcine O-type foot-and-mouth disease virus and its preparation method and application. Background technique [0002] Foot-and-mouth disease (FMD) is a severe contagious disease caused by the food and mouth disease virus (FMDV), which can cause disease and death of cloven-hoofed animals such as cattle, sheep, and pigs. The clinical symptoms of FMD are mainly manifested as the occurrence of blisters in the hooves, oral mucosa, breast skin and other places of infected animals, which in turn lead to ulceration. The most obvious clinical symptoms are high-yield dairy cows and intensively raised pigs. Although the average fatality rate of FMD is very low, the morbidity rate of infected animals is 100%, and the transmission speed is extremely fast, which leads to the reduction of...

Claims

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Application Information

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IPC IPC(8): C12N15/62C12N15/866C12N15/66C12N7/01C07K19/00A61K39/135A61P31/14
CPCY02A40/70
Inventor 陈金顶聂恺阳黄允真章洋溢杨超赵明秋
Owner SOUTH CHINA AGRI UNIV
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