Infectious pancreatic necrosis recombinant VP2 protein antigen and purification method

A protein antigen, infectious technology, applied in chemical instruments and methods, biochemical equipment and methods, peptide preparation methods, etc. Increased productivity, highly specific results

Inactive Publication Date: 2009-11-18
NORTHEAST AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there is currently no effective preventive
In terms of diagnosis and detection, virus isolation and identification of suspicious disease materials are the most intuitive diagnostic methods for this disease, but virus isolation requires certain conditions and quite a long time. Therefore, this diagnostic method is time-consuming, laborious and cannot be quickly diagnosed. Disadvantages; the most commonly used rapid diagnosis methods for this disease are serological tests, fluorescent antibody tests and enzyme-linked immunosorbent assays, etc.

Method used

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  • Infectious pancreatic necrosis recombinant VP2 protein antigen and purification method
  • Infectious pancreatic necrosis recombinant VP2 protein antigen and purification method
  • Infectious pancreatic necrosis recombinant VP2 protein antigen and purification method

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Experimental program
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Embodiment Construction

[0057] Carry out viral nucleic acid (RNA) extraction from the cell culture fluid of propagating IPNV virus, carry out the synthesis of viral genome cDNA with the downstream primer P2 of IPNV VP2 gene, use the upstream and downstream primers of IPNV VP2 gene again, carry out the amplification of VP2 gene, The amplified IPNV VP2 gene was cloned into the prokaryotic expression vector plasmid pET30b, and expressed in the Escherichia coli BL21 strain. According to the characteristics of the vector, IPTG was used in this experiment to induce the expression of the fusion protein, and the expression product was analyzed by SDS-PAGE After detecting antigenic activity with Western-blot, the fusion protein was purified using the recombinant protein expressed by the prokaryotic expression vector pET30b with 6 consecutive histidine residues, which can be combined on the Ni-NTA column, and imidazole Competitive binding on the Ni-NTA column to replace the recombinant protein provides an effec...

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Abstract

The invention provides an infectious pancreatic necrosis recombinant VP2 protein antigen and a purification method. In the invention, viral nucleic acid (RNA) is extracted from infectious pancreatic necrosis virus cell culture liquid, a downstream primer P2 of an IPNV VP2 gene is used for synthesizing a virus genome cDNA, an upstream primer and the downstream primer of the IPNV VP2 gene are utilized to amplify the VP2 gene, the IPNV VP2 gene obtained by amplification is cloned into a prokaryotic expression carrier plasmid pET30b, and expression is carried out in a colon bacillus BL21 strain to obtain the infectious pancreatic necrosis recombinant VP2 protein antigen. The antigen can overcome the potential hazards of toxin expulsion and toxin dispersion existing in an antigen in the traditional diagnosis method of infectious pancreatic necrosis, and can be used as a diagnostic antigen for disease surveillance.

Description

(1) Technical field [0001] The present invention relates to a biological preparation, and also relates to a preparation method of the biological preparation. Specifically, it relates to a fish infectious pancreatic necrosis recombinant VP2 protein antigen and a preparation method thereof. (2) Technical background [0002] Infectious pancreatic necrosis is a highly contagious and acute viral disease of salmonid fish caused by infectious pancreatic necrosis virus (IPNV). The typical symptoms are abnormal swimming and dark and dark body color of sick fish , abdominal distention, exophthalmos, hemorrhage and necrosis of subcutaneous, hepatopancreas and other visceral parenchymal organs. Eggs excrete pathogens and become a potential source of infection. Due to the widespread prevalence of the disease and the high morbidity and mortality rates, it has brought significant economic losses to the salmon and trout breeding industry in my country and the world. [0003] The traditio...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/08C12N15/62C12N15/70C07K19/00C07K1/22C12R1/19
Inventor 刘敏李一经葛俊伟唐丽杰乔薪瑗
Owner NORTHEAST AGRICULTURAL UNIVERSITY
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