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Haemophilus parasuis OppA recombinant protein antigen-containing forward haemophilus parasuis disease indirect hamagglutination diagnosis reagent and preparation method thereof

A technology of Haemophilus suis disease and Haemophilus suis, which is applied in the field of molecular biology, can solve the problems that a single antibody detection reagent cannot meet the production needs, complex antigenic components of the whole bacteria, complex serotypes, etc., to achieve strong promotion and The effect of application value, intuitive test results and strong practicability

Active Publication Date: 2013-07-10
LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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AI Technical Summary

Problems solved by technology

However, these diagnostic methods have certain limitations. The antigenic composition of the whole bacteria is complex and lacks specificity. Although the capsular polysaccharide antigen has good specificity, it can only detect a single antibody of the corresponding serotype.
However, in China, the prevalent serotypes of Haemophilus parasuis are relatively complex, mainly including serotypes 4, 5, 12, and 13. Other serotypes can occasionally occur clinically, so a single antibody detection reagent cannot meet the actual conditions. Production needs

Method used

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  • Haemophilus parasuis OppA recombinant protein antigen-containing forward haemophilus parasuis disease indirect hamagglutination diagnosis reagent and preparation method thereof
  • Haemophilus parasuis OppA recombinant protein antigen-containing forward haemophilus parasuis disease indirect hamagglutination diagnosis reagent and preparation method thereof

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Embodiment 1

[0020] 1. Haemophilus parasuis OppA Acquisition of the target gene

[0021] 1.1 Reaction system

[0022] Premixed Enzyme (Bao Bio): 12.5 , Primers (Bao Biology): 1 each , template 1 , Sterile double water: 9.5 ;

[0023] Primer: Upstream primer 5'-ATGCAAACAACCTTTACC-3'

[0024] Downstream primer 5'-TTACTGCTTAATGATATA-3';

[0025] 1.2 Amplification conditions

[0026] 94°C, 5min; 94°C, 50s; 51°C, 40s; 72°C, 100s; 35 cycles.

[0027] 2. Acquisition of OppA target protein

[0028] 2.1 will amplify OppA The target gene is connected with the cloning vector pMD19-T and transformed into Escherichia coli JM109; the positive monoclonal strain is picked and the plasmid is extracted; endonuclease EcoR I and xho I carried out double digestion of the plasmid and Pet-30a, then connected it with T4 ligase, and then transformed it into the expression strain BL21 (DE3), and induced expression with IPTG at a final concentration of 0.08mmol / L, such as figure 1 A schematic...

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Abstract

The invention discloses a haemophilus parasuis OppA recombinant protein antigen-containing forward haemophilus parasuis disease indirect hamagglutination diagnosis reagent and a preparation method thereof, wherein the reagent is fast and convenient to detect, and strong in practicability. The beneficial effects lie in that (1) the detection processes are simple and convenient, fast and accurate, and the practicability is strong, and stronger popularization and application values can be achieved; (2) the sensitivity is high, the specificity is strong and the stability is good; (3) the haemophilus parasuis 15 serotype antibody can be detected with the diagnosis reagent; and (4) the diagnosis reagent and the preparation method thereof are safe and convenient, have no need of complicated detection instruments and equipment, and the detection results are intuitive.

Description

Technical field [0001] The present invention involves molecular biology, which specifically involves the auxiliary pig's OPPA reorgan protein antigen and its coding genes, as well as the sub -pigobacteria diagnostic reagent containing the sub -pigobacteria disease containing the auxiliary pigobacterial OPPA restructuring protein antigen. Background technique [0002] Hemorrhaloboriosis of the Pig Bybolia is by the Perishalva (Hemorrhalotia ( Haemophilus parasuis HPS) Infectious diseases characterized by cellulose, multiple arthritis, and meningitis caused by cellulose.The disease occurs around the world. Pigs at 5-8 weeks are usually seen. The incidence is generally 10%-15%, and the mortality rate can reach 50%in severe cases.In recent years, the infection rate and mortality of this disease in large -scale pig farms have increased, and it has been one of the more severe bacterial diseases that endanger the pig industry. [0003] Hemodolisobacteria is a conditional disease of worl...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/285C12N15/31C12N15/63C12N5/10C12N1/21C12N15/70G01N33/68C12R1/21
Inventor 逯忠新高鹏程贺英储岳峰赵萍
Owner LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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