Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Pseudorabies virus epitope polypeptide gene engineering vaccine

A technology of epitope polypeptide and pseudorabies, applied in the field of biotechnology genetic engineering

Active Publication Date: 2015-05-20
QINGDAO MINGQIN BIOLOGICAL TECH CO LTD
View PDF2 Cites 8 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Monoclonal antibodies to gB, gC, and gD passively protect mice and pigs against lethal PRV infection, while gE mAbs only protect mice

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Pseudorabies virus epitope polypeptide gene engineering vaccine
  • Pseudorabies virus epitope polypeptide gene engineering vaccine
  • Pseudorabies virus epitope polypeptide gene engineering vaccine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0045] Example 1 Design idea of ​​pseudorabies epitope polypeptide vaccine protein

[0046] According to the amino acid sequences of glycoproteins gB, gC and gD of the main epidemic strains of pseudorabies in China, the present invention utilizes relevant bioinformatics software DNASTAR, BIMAS and SYFPEITHI to carry out hydrophilicity, antigenicity, plasticity, surface accessibility and Garnier- Robson's secondary structure was analyzed, and then oligonucleotide fragments were designed according to the conservation of B cell neutralizing epitope and T cell epitope position and amino acid sequence, and bovine herpes virus type I envelope protein VP22 was introduced as an adjuvant molecule. The designed pseudorabies virus B cell and T cell epitopes and BVP22 molecular polypeptides were co-expressed in Escherichia coli in series, and after fermentation, purification, emulsification and other processes, a pseudorabies epitope polypeptide vaccine with ideal immunogenicity was obtain...

Embodiment 2

[0049] Example 2 Construction of Escherichia coli expression vector and expression strain

[0050]The nucleotide encoding the polypeptide designed in Example 1 was sent to Shanghai Handsome Biotechnology Co., Ltd. for synthesis, and BamH I (5' end) and HindIII (3' end) restriction enzyme sites were designed at both ends of the nucleotide fragment After the two fragments were synthesized, they were respectively cloned into the pMD18T vector, and sequence determination confirmed that the inserted gene fragment was consistent with the designed sequence (see the sequence listing). The recombinant plasmid was named pMD18T-PRV(gB / gC / gD)-BVP22, and the two plasmids were digested with corresponding restriction enzymes. The E. coli expression vector was pRSETB plasmid from Invitrogen Company, and the same Restriction endonuclease treatment, digestion conditions: 10 μl reaction system, 2 μl plasmid was added to the system, the restriction enzyme was 5 active units (New England biolabs),...

Embodiment 3

[0054] Example 3 Fermentation, purification and emulsification of engineering bacteria

[0055] The production strains were taken for fermentation, inoculated into 2ml LB liquid medium (containing 100 μg / ml ampicillin), and cultured at 37° C. with shaking at 180 rpm 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....

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention relates to preparation and application of a Pseudorabies virus (PRV) epitope polypeptide gene engineering vaccine. The vaccine uses PRV as the main glycoprotein, and B cell neutralizing epitope of gB, gC and gD and T cell immune epitope as the vaccine frame structure. The vaccine is prepared by the following steps: connecting the main glycoprotein with the vaccine frame structure through a flexible linker, connecting in series with a molecular adjuvant Bovine Herpesvirus 1 (BHV-1) envelope protein VP22, cloning a pRSETB vector, transforming Escherichia coli, fermenting, purifying, emulsifying and the like to obtain the PRV epitope polypeptide gene engineering vaccine. The invention also relates to an application method of the vaccine. The animal experiment indicates that the PRV epitope polypeptide gene engineering vaccine has equivalent effect to the live virus vaccine in the aspect of humoral immunity and cellular immunity level, and can be stimulated to generate T lymphopoiesis immunoreaction on the cellular level and generate antibody immunoreaction with virus neutralizing activity on the humoral level.

Description

technical field [0001] The invention belongs to the field of biotechnology genetic engineering, and mainly relates to the preparation and application of a pseudorabies (Pseudorabies virus, PrV) epitope polypeptide genetic engineering vaccine. Specifically, using genetic recombination technology, the B cell neutralizing epitope and T cell immune epitope of the main glycoproteins gB, gC, and gD of PRV are connected in series with the bovine herpesvirus type 1 (Bovine Herpesvirus 1, BHV-1) envelope protein VP22 , and cloned into a vector, transformed into a host bacterium, prepared through fermentation, purification and emulsification processes, to obtain a pseudorabies epitope polypeptide genetic engineering vaccine and the application of the vaccine in preventing pseudorabies, a major animal disease. Background technique [0002] Pseudorabies is an acute infectious disease that occurs in domestic animals, wild mammals, companion animals and experimental animals, with fever, i...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): C07K19/00C12N15/62C12N15/70C12N1/21A61K39/245A61P31/22
Inventor 李殿明蒲勤张毓金齐春梅田春辉刘甜甜任百亮张导春党将将吴启凡
Owner QINGDAO MINGQIN BIOLOGICAL TECH CO LTD
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com