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Application of antigen causing antibody-dependent enhancement of Tembusu virus

A Tembusu virus and antibody-dependent technology, applied in the fields of molecular biology and immunology, can solve the problems of unsatisfactory protection effect and enhanced pathogenicity, and achieve the effect of ensuring protection effect and promoting virus proliferation

Active Publication Date: 2018-10-19
JIANGSU ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, due to the antibody-dependent enhancement of Tembusu virus, the protective effect of the vaccine after immunization is not ideal, and there is a hidden danger of promoting virus proliferation and enhancing pathogenicity after secondary infection

Method used

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  • Application of antigen causing antibody-dependent enhancement of Tembusu virus
  • Application of antigen causing antibody-dependent enhancement of Tembusu virus

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] Example 1 Acquisition of Antigens that Cause Tambusu Virus Antibody-Dependent Enhancement

[0021] According to the coding gene sequence of the antigen that caused the Tembusu virus antibody-dependent enhancement, the coding gene fragment was obtained by artificial synthesis, and inserted into the plasmid pGEX-4t-1 through the EcoR I and Sal I restriction sites to obtain the recombinant plasmid pGEX-4t-1. 4t-28. The expression vector pGEX-4t-28 was introduced into Escherichia coli to obtain an engineered strain expressing the antigen. Inoculate the LB medium with the engineering strain obtained, and wait for the OD 600nm When the concentration is 0.4-0.6, add IPTG with a final concentration of 1 mmol / L to induce protein expression in engineering bacteria containing the expression vector pGEX-4t-B. The expression product was purified using Glutathione-agarose (Sigma), and the operation steps were performed according to the instructions. The purified antigen was identi...

Embodiment 2

[0022] Example 2 The antigen-induced production of specific antibodies caused by the dependent enhancement of Tembusu virus antibodies

[0023] 1. The antigenic protein obtained in Example 1 was used to immunize 9-day-old ducklings, 200 μg each, emulsified with an equal volume of Freund's complete adjuvant for the first immunization. Two weeks after the immunization, a booster immunization was carried out, 200 μg per mouse, and the booster immunization was emulsified with an equal volume of Freund's incomplete adjuvant. The ducklings in the control group were immunized with the same volume of PBS, and the other treatments were the same.

[0024] 2. Two weeks after the booster immunization, the ducklings in the immunized group and the control group were subjected to wing vein blood collection, the serum was separated, and the specific antibody titer was detected by indirect ELISA.

[0025] Implementation results: the titers of specific antibodies in the serum of ducklings in t...

Embodiment 3

[0026] Example 3 Promoting effect of antigen-specific antibody on virus proliferation in vivo

[0027] 1. The ducklings after 2 weeks of booster immunization in Example 2 were subjected to the Tembusu virus challenge test. Each attack dose is 10 4 TCID 50 .

[0028] 2. 1-5 days after the challenge, the venous blood of the young duck wings was aseptically collected every day, the serum was separated, and the BHK-21 cells that had grown into a monolayer were inoculated. After incubation at 37°C for 2h, wash with PBS three times, add RPMI1640 containing 1% fetal bovine serum, and culture at 37°C for 3 days. The cells were collected, and the viral nucleic acid content was detected by fluorescent quantitative RT-PCR. Using the relative quantification method (2 -ΔΔCt ) for analysis. The primers used to detect viral nucleic acid are: upstream primer: 5'-GTGAGATCTTACTGCTATGAG-3'; downstream primer: 5'-ACTTGGCACATGTCTGTATGC-3'. GAPDH gene was used as internal reference, upstream...

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Abstract

The present invention discloses application of an antigen causing antibody-dependent enhancement of Tembusu virus. The antigen causing the antibody-dependent enhancement of Tembusu virus has an aminoacid sequence shown in SEQ ID NO.1. The antigen causing the antibody-dependent enhancement of the Tembusu virus is applied to preparation of a Tembusu virus vaccine. According to the invention, the epitope protein expressed by escherichia coli is used for immunizing ducklings, so that high-level specific antibodies can be induced in the ducklings, and the antibodies can promote virus proliferationand increase the titer of the virus in the ducklings. In the future research and development of the Tembusu virus vaccine, this kind of antigen can be removed through molecular design to avoid the antibody-dependent enhancement caused by the vaccine and ensure the protection effect. Therefore, identifying the antigen causing the antibody-dependent enhancement of the Tembusu virus has an importantapplication value in developing effective and safe vaccines and preventing and controlling the epidemic of the Tembusu virus.

Description

technical field [0001] The invention belongs to the technical fields of molecular biology and immunology, and relates to the application of an antigen that induces the antibody-dependent enhancement of Tambusu virus. Background technique [0002] Tambusu virus disease is a new disease characterized by a sharp loss of appetite and a sudden drop in egg production, which has caused significant economic losses to the duck and goose farming industry. The flavivirus genus includes dengue virus, Japanese encephalitis virus, West Nile virus, yellow fever virus, Zika virus and other arboviruses. [0003] Viruses initiate infection by attaching surface proteins to specific receptors on host cells. Antibodies specific to virus surface proteins can "neutralize" the virus by blocking the surface protein and inhibiting the adhesion of the virus to the receptor, making it lose the ability to infect cells. In some cases, however, antibodies play the opposite role during viral infection. ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/18C12N15/40C12Q1/70C12Q1/686A61K39/12A61P31/14G01N33/569
CPCA61K39/12A61K2039/552A61P31/14C07K14/005C12N2770/24122C12N2770/24134C12Q1/686C12Q1/701G01N33/56983G01N2333/185
Inventor 赵冬敏章丽娇韩凯凯黄欣梅杨婧刘宇卓刘青涛李银
Owner JIANGSU ACAD OF AGRI SCI
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