Recombination Lactococcus lactics for expressing pig infectious gastroenteritis virus S protein and preparation method thereof

A kind of Lactococcus lactis, infectious technology, applied in the field of preparation of biological products, can solve the problem of unsatisfactory immune protection effect

Inactive Publication Date: 2007-10-24
NORTHEAST AGRICULTURAL UNIVERSITY
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  • Abstract
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Problems solved by technology

Practice has proved that the secretory antibody (sIgA) produced by intestinal mucosal immunization is an effective antibody against TGEV infec

Method used

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  • Recombination Lactococcus lactics for expressing pig infectious gastroenteritis virus S protein and preparation method thereof
  • Recombination Lactococcus lactics for expressing pig infectious gastroenteritis virus S protein and preparation method thereof
  • Recombination Lactococcus lactics for expressing pig infectious gastroenteritis virus S protein and preparation method thereof

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Embodiment Construction

[0022] The present invention is described in more detail below in conjunction with accompanying drawing example:

[0023] 1. Construction of TGEV S gene Lactococcus lactis expression vector

[0024] Using the plasmid pUC-S as a template, using TS7 and TS8 as primers to amplify the fragment containing the main antigenic sites of TGEV S gene A, B, C and D, and named it Sa, the Sa gene was digested with the pNZ8112-Sa vector plasmid, Connected and transformed into competent cell NZ9000 by electrotransformation, the Lactococcus lactis containing the recombinant plasmid was named pNZ8112-Sa / NZ9000.

[0025] 2. Induced expression of TGEV S protein in Lactococcus lactis

[0026] The pNZ8112-Sa / NZ9000 positive recombinant bacteria and the pNZ8112 / NZ9000 empty plasmid control bacteria were inoculated in GM17 liquid medium, and after anaerobic culture at 30°C overnight, the overnight culture was inoculated with a volume ratio of 1:25 for amplification and culture in In 20ml culture me...

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Abstract

The invention provides a recombination lactic acid galactococcus method of expessing pig transmissible gastroenteristis virus mutation S protein, which comprises the following steps: designing double primers according to the total gene order of pig Transmissible gastroenteristis virus S protein and gene fusion characteristic of expressing the carrier plasmid; proceeding PCR; getting the 2007bp destination fragments of four main antigen sites with TGEV S gene; connecting with the carrier plasmid PNZ8112 of secretory expression; getting into the host lactic acid galactococcus NZ9000 cell by electric conversion; expressing under the inducing of nisin. The invention constructs the expression carrier system of coronaviridae TGEV S mutein lactic acid galactococcus, which expresses about 66KDa destination protein of four main antigen sites with TGEV. Immunological ink mist experiment and indirect immunological fluorescein test indicate that the exogenesis protein is able to react with TGEV immunological serum and the recombination S protein has the same antigenic as TGEV natural antigenic.

Description

(1) Technical field [0001] The present invention relates to a biological product, and the present invention also relates to a preparation method of the biological product. (2) Background technology [0002] Porcine transmissible gastroenteritis (TGE) is a digestive tract infectious disease characterized by piglet vomiting, severe diarrhea and high lethality caused by porcine transmissible gastroenteritis virus (TGEV) of the Coronaviridae family. . The disease is one of the important diseases of early death of piglets in my country and various pig-raising countries in the world. Vaccination is the main measure to prevent this disease. Practice has proved that the secretory antibody (sIgA) produced by intestinal mucosal immunization is an effective antibody against TGEV infection, while other serum antibodies produced by oral immunization, such as IgG and IgM, are not ideal for the immune protection of this disease . Therefore, it is of great significance to choose a carri...

Claims

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Application Information

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IPC IPC(8): C12N1/21C12N15/33C12N15/09C07H21/04C12N15/74G01N33/53
Inventor 唐丽杰李一经葛俊伟欧笛
Owner NORTHEAST AGRICULTURAL UNIVERSITY
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