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Construction method of PRV gE/gI dual-gene deletion strains for expressing ASFV P30 protein

A construction method and dual-gene technology, which can be applied in genetic engineering, plant genetic improvement, chemical instruments and methods, etc., can solve the problems of pig farming and the spread of ASFV, and achieve low cost, prevention of ASFV and PRV infection, Prepare simple effects

Inactive Publication Date: 2020-07-31
SICHUAN AGRI UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

So far, there is no effective vaccine in the world, and the spread of ASFV will bring a devastating blow to the pig industry

Method used

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  • Construction method of PRV gE/gI dual-gene deletion strains for expressing ASFV P30 protein
  • Construction method of PRV gE/gI dual-gene deletion strains for expressing ASFV P30 protein

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Embodiment Construction

[0035] The following will clearly and completely describe the technical solutions in the embodiments of the present invention with reference to the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only some, not all, embodiments of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by persons of ordinary skill in the art without creative efforts fall within the protection scope of the present invention.

[0036] The present invention provides a technical solution: a method for constructing a PRV gE / gI double gene deletion strain expressing ASFV P30 protein, specifically comprising the following steps,

[0037] S1: Reagent preparation:

[0038] LB liquid medium; fast restriction enzymes (Xho I, Sal I); Lipefectamine TM 3000 transfection reagent; endotoxin-free plasmid mini-extraction kit; DH5α Escherichia coli competent cells;

[0039] LB liquid medium is obtained b...

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Abstract

The invention discloses a construction method of PRV gE / gI dual-gene deletion strains for expressing ASFV P30 protein and belongs to the biopharmaceutical field. The construction method specially comprises the following steps of S1, preparing a reagent; S2, preparing cloned plasmids; S3, constructing a PRV eukaryon transfer plasmid pEGFP-CP204L; S4, performing cotransfection on DNA of PRV and pEGFP-CP204L plasmids. P30 protein gene (CP204L) fragments are synthesized, the transfer plasmid pEGFP-CP204L is constructed, a classical homologous recombination manner is adopted for inserting an ASFV CP204L gene into a PRV genome to replace gI and gE virulence genes, and recombinant pseudorabies virus strains rXJgE / gI-CP204L for constructing co-expression P30 protein can be used for preventing ASFVand PRV infection, and the cloned plasmids are simple to prepare, low in cost and suitable for industrial production.

Description

technical field [0001] The invention relates to the technical field of biomedicine, in particular to a method for constructing a PRV gE / gI double gene deletion strain expressing ASFV P30 protein. Background technique [0002] African swine fever (African swine fever, ASF) is an acute, febrile, highly contact and fatal zoonotic disease caused by African swine fever virus (ASFV). In addition to being able to infect pigs of all breeds and ages, African swine fever manifests as symptoms such as high fever, systemic hemorrhage, respiratory disorders and neurological symptoms. Due to the difference in virulence of African swine fever virus, the infection process and clinical symptoms of African swine fever are significantly different, and the most acute and acute infection fatality rates are as high as 100%. So far, there is no effective vaccine in the world, and the spread of ASFV will bring a devastating blow to the pig industry. On August 3, 2018, the African swine fever epid...

Claims

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Application Information

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IPC IPC(8): C12N7/01C12N15/34C12N15/85C12N15/66A61P31/20A61P31/22
CPCA61P31/20A61P31/22C07K14/005C12N7/00C12N15/66C12N15/85C12N2710/12022C12N2710/12034C12N2710/16721C12N2710/16722C12N2710/16734
Inventor 徐志文朱玲曾喻兵
Owner SICHUAN AGRI UNIV
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