Construction method and application of recombinant porcine reproductive and respiratory syndrome virus capable of expressing African swine fever virus p12 or p17 proteins

An African swine fever virus, respiratory syndrome technology, applied in the field of bioengineering, can solve the problems of genetic instability, inability to function, loss of coding sequences, etc.

Inactive Publication Date: 2020-03-24
SHANGHAI VETERINARY RES INST CHINESE ACAD OF AGRI SCI
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  • Abstract
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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, showing genetic instability

Method used

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  • Construction method and application of recombinant porcine reproductive and respiratory syndrome virus capable of expressing African swine fever virus p12 or p17 proteins
  • Construction method and application of recombinant porcine reproductive and respiratory syndrome virus capable of expressing African swine fever virus p12 or p17 proteins
  • Construction method and application of recombinant porcine reproductive and respiratory syndrome virus capable of expressing African swine fever virus p12 or p17 proteins

Examples

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Embodiment 1

[0031] Example 1 Construction of recombinant PRRSV plasmid expressing ASFV p12 or p17 protein

[0032] According to the nucleotide sequences of GenBank accession numbers EF635006 and MH766894, 6 primers were designed for SOE PCR amplification. The process is as follows figure 2 As shown, cell transfection experiments were performed to verify the infectivity of the obtained chimeric recombinant plasmids. The specific process is as follows:

[0033] 1.1 Primer design

[0034] According to the base sequence of HuN4 on GenBank and the base sequence of the popular type II ASFV genome in my country, two sets of SOE PCR primers are designed. The first set of SOE PCR primers are named: HF11559, HR13090, ASFV-p12-F2, ASFV- The sequences of p12-R1, ASFV-p12-F4, ASFV-p12-R2 / R3 are as follows:

[0035] HF11559: 5’-TCATACATCCGAGTTCCTGTT-3’ (SEQ ID NO.1)

[0036] HR13090: 5’-GAAATATTGTCATGGCGAGGC-3’ (SEQ ID NO. 2)

[0037] ASFV-p12-F2: 5’-TCATTGAACCAACTTTAGGCCTGAATTGAAatggcacttgatggttcaagtgg -3’ (SEQ...

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Abstract

The invention provides a construction method of porcine reproductive and respiratory syndrome virus (PRRSV) recombinant plasmid for expressing ASFV (African swine fever virus) p12 protein or p17 protein. The invention also provides a genetic engineering vaccine constructed according to the recombinant plasmid and a construction method. Indirect immunofluorescence is carried out on rPRRSV-p12 and rPRRSV-p17 infection holes, and it is found that specific fluorescence for resisting PRRSV N protein and specific fluorescence for resisting ASFV p12 protein or p17 protein appear in the visual field.It shows that the recombinant PRRSV expressing African swine fever virus p12 protein or p17 protein can ensure that the p12 or p17 protein of exogenous protein ASFV can be well, efficiently and stablyexpressed. Researches show that p12 and p17 are important antigen proteins and can be used for ASF vaccine development.

Description

Technical field [0001] The present invention belongs to the field of bioengineering. Specifically, it relates to a virus recombinant plasmid and genetic engineering vaccine, and more specifically, to a PRRS virus recombinant plasmid capable of expressing African swine fever virus (ASFV) p12 or p17 protein and Genetically engineered vaccine. Background technique [0002] ASFV is an acute, severe, hemorrhagic, and highly contagious infectious disease caused by ASFV infection of domestic pigs or wild boars. The course of the disease is short, and the morbidity and mortality are high. A serious threat to the global pig industry. There are no effective vaccines and treatments. ASF first appeared in Kenya, Africa in 1921 and spread to many regions in Africa; it spread to Portugal in Europe in 1957; it was introduced to Cuba in South America from Western Europe in 1971, followed by outbreaks in South American countries; it was introduced to Europe and Asia in 2007 The Caucasus regio...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/85C12N15/34C12N15/40A61K39/12A61P31/14A61P31/20
CPCC12N15/85C07K14/005A61K39/12A61P31/14A61P31/20C12N2770/10022C12N2770/10034C12N2710/12022C12N2710/12034A61K2039/53A61K2039/552A61K2039/70
Inventor 童光志高飞李国新童武姜一峰郑浩周艳君虞凌雪李丽薇刘长龙
Owner SHANGHAI VETERINARY RES INST CHINESE ACAD OF AGRI SCI
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