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Porcine reproductive and respiratory syndrome recombinant adenovirus rAd-GF35

A technology for recombinant adenovirus and respiratory syndrome, which is applied in the direction of antiviral agents, viral antigen components, viruses/bacteriophages, etc., to achieve the effect of broad application prospects

Inactive Publication Date: 2009-05-06
NANJING AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

GMCSF has a good adjuvant effect and is widely used in antiviral and antitumor research. At present, there is no report on the application of GMCSF to the recombinant adenovirus constructed into porcine reproductive and respiratory syndrome virus PRRSV

Method used

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  • Porcine reproductive and respiratory syndrome recombinant adenovirus rAd-GF35
  • Porcine reproductive and respiratory syndrome recombinant adenovirus rAd-GF35
  • Porcine reproductive and respiratory syndrome recombinant adenovirus rAd-GF35

Examples

Experimental program
Comparison scheme
Effect test

Embodiment Construction

[0043] (1) Construction of Genetic Engineering Body—PRRSV Recombinant Adenovirus (rAd-GF35)

[0044] 1 Amplification, cloning, identification and sequencing analysis of the target gene

[0045] 1.1 Primer design

[0046] Seven primers were designed with reference to the gene sequence of PRRSV S Y0608 strain (GenBank Accession Number EU144079) and the porcine GMCSF gene sequence of accession number U67175:

[0047] SY-GP3.1: 5-GACCTCGAGATGGCTAATAGCTGTACATT-3;

[0048] SY-GP3.2: 5-ACAAAGCTTTCGCCGTGGGCACT-3;

[0049] SY-GP5.1:5-ACGAAGCTTATGTTGGGGAAGTGCT-3;

[0050] SYGP5.2: 5-ACGAAGCTTATGTTGGGGAAGTGCT-3;

[0051] GMCSF-P1: 5-ACGAAGCTTATGTTGGGGAAGTGCT-3;

[0052] GMCSF-2AP2.1s: 5-GACGTCGCCGGCCAACTTGAGAAGGTCAAAGTTCTTTTTGACTGGCCCC-3;

[0053] GMCSF-2AP2.2s: 5-GCACTCGAGGGGCCCTGGGTTGGACTCGACGTCGCCGGCCAACTTGAG-3

[0054] After stimulating porcine peripheral blood mononuclear cells with concanavalin ConA (5ug / ml), according to TRIzol (Invitrogen) reagent The manual extracts the...

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Abstract

The invention discloses a virus (PRRSV) recombinant adenovirus rAd-GF35 of porcine reproduction and respiratory syndrome, which belongs to the technical field of high-tech biotechnology. Highly pathogenic PRRSV SY0608 separation strains of GP3 and GP5 protein genes and porcine colony sell stimulating factor (GMCSF) whole gene sequences are amplified, connected in series and cloned into pShuttle-CMV to be transformed into escherichia coli BJ5183 together with pAdEasy-1 so as to obtain recombinant plasmids; and HEK293A cells are transfected to obtain the recombinant adenovirus so as to successfully express PRRSV SY0608 strains GP3 and GP5 and a GMCSF protein. The recombinant virus can express the GMCSF correctly, and play a role of an adjuvant so as to improve the immune activity of the PRRSV GP3 and GP5 proteins and stimulate the immune protective reaction of an organism more effectively. The recombinant adenovirus vaccine has the safety of a subunit vaccine and the antigen proliferating ability of an attenuated vaccine, thereby having wide development and application prospect.

Description

1. Technical field [0001] The invention relates to porcine reproductive and respiratory syndrome virus (PRRSV) recombinant adenovirus rAd-GF35 and a vaccine, which belongs to the high-tech field of biotechnology and is specially used to strengthen the immune protection of pigs against porcine reproductive and respiratory syndrome virus and reduce the risk of pig farms Economic losses. 2. Background technology [0002] Recombinant adenovirus genetic engineering technology has been widely used in human gene therapy, and many recombinant adenoviruses used in human gene therapy have entered the third phase of clinical trials. Because the recombinant adenovirus has a wide range of hosts, can infect many types of cells, can express human and non-human proteins, and will not insert into the host chromosome, has a low reverse mutation rate, and can be replicated to a high titer. The GP3 GP5 protein is a glycoprotein of about 41 kD and 25 kD encoded by ORF3 ORF5 of porcine reproduct...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/01C12N15/40C12N15/861A61K39/12A61P31/14
Inventor 李玉峰王兴龙姜平
Owner NANJING AGRICULTURAL UNIVERSITY
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