Pig viral infectious disease gene recombined live vaccine using canine II type adenovirus as carrier and preparation process thereof

A production process and gene recombination technology, applied in genetic engineering, gene therapy, antiviral agents, etc., to achieve the effects of long shelf life, good immune protection effect, and good genetic stability

Inactive Publication Date: 2006-09-06
MILITARY VETERINARY RES INST PLA MILITARY MEDICAL ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

But these vaccines all have the disadvantages of traditional vaccines

Method used

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  • Pig viral infectious disease gene recombined live vaccine using canine II type adenovirus as carrier and preparation process thereof
  • Pig viral infectious disease gene recombined live vaccine using canine II type adenovirus as carrier and preparation process thereof
  • Pig viral infectious disease gene recombined live vaccine using canine II type adenovirus as carrier and preparation process thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037]The preparation of embodiment 1 recombinant swine influenza virus HA gene adenovirus vector live vaccine

[0038] According to the process route shown in Figure 1, the recombinant adenovirus live vector vaccine was prepared. Under the electron microscope, the recombinant virus had the general characteristics of adenovirus. After infecting MDCK cells, specific cytopathic changes could occur. See attached drawing 4. Specific steps are as follows:

[0039] 1. The rescue plasmid pPoly2-UD containing the paired ends of the CAV-2 genome was obtained by PCR method, which was linearized with BstB I+Hind III enzymes and then homologously recombined with the CAV-2 genomic DNA to obtain the CAV-2 genome-wide cloning plasmid pPoly2 -CAV / 2.

[0040] PCR primers:

[0041] P 1 : 5′ end GCG-TGC-CTA-ACT-ACA-AAC-TCA-AT

[0042] 5' end G-CCT-GCT-GCT-TGA-TTT-TGT-GAT-C

[0043] P 2 : 5′ end A-CTC-ATA-GAA-GTA-GGC-AGC-TCC-G

[0044] 5' end CAT-CAT-CAA-TAA-TAT-ACA-GGA-CAA-AG

[0...

Embodiment 2

[0065] Example 2 Preparation of recombinant classical swine fever virus E2 gene adenovirus vector live vaccine

[0066] According to attached image 3 The process route shown is to prepare the recombinant adenovirus live vector vaccine. The recombinant virus has the general characteristics of adenovirus under the electron microscope. After infecting MDCK cells, specific cytopathic changes may occur. See attached drawing 4. Specific steps are as follows:

[0067] 1. The rescue plasmid pPoly2-UD containing the paired ends of the CAV-2 genome was obtained by PCR method, which was digested with PstI+Hind III and linearized, and then homologously recombined with the CAV-2 genomic DNA to obtain the CAV-2 whole genome cloning plasmid pPoly2- CAV / 2.

[0068] PCR primers:

[0069] P 1 : 5′ end GCG-TGC-CTA-ACT-ACA-AAC-TCA-AT

[0070] 5' end G-CCT-GCT-GCT-TGA-TTT-TGT-GAT-C

[0071] P 2 : 5′ end A-CTC-ATA-GAA-GTA-GGC-AGC-TCC-G

[0072] 5' end CAT-CAT-CAA-TAA-TAT-ACA-GGA-CAA-...

Embodiment 3

[0092] Embodiment 3 Preparation of recombinant porcine AsiaI type foot-and-mouth disease virus VPI gene adenovirus vector live vaccine

[0093] According to attached image 3 The process route shown is to prepare the recombinant adenovirus live vector vaccine. The recombinant virus has the general characteristics of adenovirus under the electron microscope. After infecting MDCK cells, specific cytopathic changes may occur. See attached drawing 4. Specific steps are as follows:

[0094] 1. Obtain the CAV-2 whole genome cloning plasmid pPoly2-CAV2 by the same method as described in Examples 1, 2, and 3.

[0095] 2. The PCR method obtains VPI from the AsiaI type foot-and-mouth disease virus PI whole gene, and the PCR primers are:

[0096] Upstream primer: 5'-CCGGAATTCGCCATGGCTCGCCGGCAGACTACCAC-3'

[0097] Downstream primer: 5'-CCGCTCGAGTTATAGCCTGTTTCTCGGGTGCAA-3' Restriction sites EcoRI and XhoI were added to the upstream and downstream primers respectively.

[0098] 3. Diges...

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Abstract

This invention supplies a series of production techniques of gene recombination live vaccine of swine virus contagion with canine ó� adenovirus as carrier and finished goods. The viral live vectors vaccine takes swine important virus zymad protective antigens gene as object gene, which are chosen from HCV-E1, E2 gene, FMDV-VP1íóVP2íóVP3íóVP4 gene, TGEV-SíóNíóM gene, PEDV-SíóNíóM geneú¼SIV-HAíóNA geneú¼RV-GíóN gene, etc. The produced vaccines contain recombined swine influenza virus HA gene adenovirus carrier live vaccine, swine plague virus E2 gene adenovirus carrier live vaccine and recombination swine AsiaI foot-and-mouth disease virus VPI gene adenovirus carrier live vaccine. Recombination virus has good inheritance stability, and vaccine immunization can induct pig develop differential antiviral neutralization antibody. It has good immune protection effect and has no toxic side effect. The goods are facilitating for preserve and transportú”it has long storage life and simple technics, and it fits for commercial manufacture.

Description

Technical field: [0001] The present invention aims to provide a series of gene recombined live vaccines and production techniques for porcine viral infectious disease immunity with canine type II adenovirus as the carrier, which are used in the development and production of porcine viral infectious disease vaccines to prevent porcine viral infection It belongs to the technical field of prevention and treatment of animal infectious diseases. Background technique: [0002] Viral infectious diseases of pigs such as swine fever, influenza, rabies, foot-and-mouth disease, porcine transmissible gastroenteritis, and porcine epidemic diarrhea not only seriously endanger the development of the pig industry, but also cause huge economic losses every year, and swine flu, rabies, etc. The occurrence and prevalence of zoonotic diseases also seriously threaten human life and health, so the prevention and control of these diseases has become the focus of veterinary work and research hotspo...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61K39/145A61K39/135A61K9/19C12N15/861A61P31/12
Inventor 夏咸柱扈荣良张守峰高玉伟杨松涛王承宇
Owner MILITARY VETERINARY RES INST PLA MILITARY MEDICAL ACAD OF SCI
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