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Construction method and application of recombinant prrs virus genetic engineering vaccine expressing classical swine fever virus e2 protein

A gene and virus technology, applied in the fields of PRRS virus recombinant plasmids and genetic engineering vaccines, can solve problems such as failure to function, loss of coding sequences, genetic instability, etc., and achieve good immune effects and good safety effects

Active Publication Date: 2018-12-11
SHANGHAI VETERINARY RES INST CHINESE ACAD OF AGRI SCI
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

But even so, the insertion of foreign genes will gradually lose the coding sequence in the process of passage, so that it cannot perform its function
genetic instability

Method used

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  • Construction method and application of recombinant prrs virus genetic engineering vaccine expressing classical swine fever virus e2 protein
  • Construction method and application of recombinant prrs virus genetic engineering vaccine expressing classical swine fever virus e2 protein
  • Construction method and application of recombinant prrs virus genetic engineering vaccine expressing classical swine fever virus e2 protein

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Embodiment 1

[0034] The construction of the chimeric recombinant plasmid that embodiment 1 expresses CSFV E2 protein

[0035] According to the base sequences of GenBank accession numbers EF635006 and HM175885, 6 primers were designed for SOE PCR amplification. The process is shown in Figure 2. Then, the obtained chimeric recombinant plasmids were verified by cell transfection experiments. Infectious, the specific process is as follows:

[0036] 1.1 Primer design

[0037] According to the base sequence of HuN4-F112 with GenBank accession number EF635006 and the sequence of swine fever virus with accession number HM175885, SOE PCR primers were designed and named as: HF11559, HR13090, CSFV-E2-F2, CSFV-E2-R1, The sequences of CSFV-E2-F4 and CSFV-E2-R2 / R3 are as follows:

[0038] HF11559: 5'-TCATACATCCGAGTTCCTGTT-3' SEQ ID NO.1

[0039] HR13090: 5′-GAAATATTGTCATGGCGAGGC-3′SEQ ID NO.2

[0040] CSFV-E2-F2: 5'-TCATTGAACCAACTTTAGGCCTGAATTGAAatggcattcctcatctgcttg-3'SEQ ID NO.3

[0041] CSFV-E2-...

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Abstract

The invention provides a method for constructing recombinant plasmids of porcine reproductive and respiratory syndrome virus (PRRSV) capable of expressing classic swine fever virus (CSFV) E2 protein, and a genetically engineered vaccine constructed according to the recombinant plasmids. Based on a reverse genetic operating platform, pA-SM-E2 and pA-C-E2 recombinant plasmids are constructed according to E2 genes of CSFV Shimen strains and hog cholera virus strains C, and genes of highly pathogenic porcine reproductive and respiratory syndrome virus cell attenuated vaccine strains HuN4-F112. After MARC-145 cells are transfected, live viruses are successfully rescued, and a situation that recombinant viruses (vA-SM-E2 and vA-C-E2) and a parental virus vHuN4-F112 have similar virological characteristics is found. After the vA-SM-E2 is selected for carrying out an immunological experiment on porcine bodies, the vA-SM-E2 has good security to the porcine bodies, and the vA-SM-E2 after being immunized can induce the bodies to produce immune response so as to produce high-level PRRSVN protein and CSFV E2 protein based antibodies, therefore, the vA-SM-E2 is a candidate strain of the genetically engineered vaccine.

Description

Background technique: [0001] The invention belongs to the field of bioengineering, and in particular relates to a virus recombinant plasmid and a genetic engineering vaccine, and more specifically relates to a PRRS virus recombinant plasmid and a genetic engineering vaccine capable of expressing CSFV E2 protein. [0002] Porcine reproductive and respiratory syndrome (Porcine Reproductive and Respiratory Syndrome, PRRS) is a contagious infectious disease that seriously affects the pig industry, causing huge economic losses to countries all over the world. The pathogen PRRSV has been in the process of continuous mutation and evolution. At present, reverse genetic manipulation is widely used to study the biological characteristics, pathogenic mechanism, virulence determinants and molecular mechanism of continuous variation of PRRSV, and the study of its genome as an exogenous gene expression vector is also widely carried out. Except for ORF1 and ORF2 and ORF4 and ORF5 in the PRR...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/85C12N15/66A61K39/295A61K39/187A61K39/12A61P31/14
Inventor 童光志高飞姜一峰李国新周艳君虞凌雪李丽薇童武郑浩杨莘
Owner SHANGHAI VETERINARY RES INST CHINESE ACAD OF AGRI SCI
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