Goat contagious pleuropneumonia subunit vaccine as well as preparation method and application thereof

A subunit vaccine and pleuropneumonia technology, applied in the field of animal infectious disease vaccine preparation, can solve the problems of limited protective effect, reduce immune side reactions, provide immune protection effect, and improve immune response

Active Publication Date: 2021-06-01
LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Mycoplasma goat pneumoniae subspecies immune proteins A, B, C, D and E related to the present invention are analyzed on the basis of Mccp whole genome, comparative genomics, proteomics, immunoproteomics, etc., to screen immunogenicity Further comprehensive analysis of protein cell localization, protein hydrophilicity, antigenicity, signal peptide, transmembrane region, T cell epitope, B cell epitope, etc., and through in vivo and in vitro tests to verify and classify and screen; the above immune protein combinations can overcome single Proteins are difficult to induce comprehensive humoral and cellular immunity, and the protective effect is limited. Screen the immune protein combination and add an adjuvant formula. The prepared subunit vaccine can induce a comprehensive immune response and provide good protection against Mccp infection in goats.

Method used

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  • Goat contagious pleuropneumonia subunit vaccine as well as preparation method and application thereof
  • Goat contagious pleuropneumonia subunit vaccine as well as preparation method and application thereof
  • Goat contagious pleuropneumonia subunit vaccine as well as preparation method and application thereof

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preparation example Construction

[0043]The present invention also provides the preparation method of the goat infectious pleuropneuman sub-unit vaccine, including the following steps: 1) combination of the goats pneumonia in the goat pneumonia and saponin in the goats and saponin, and the antigen saponin mixture is allowed. The static temperature is 2 to 8 ° C, the standing time is 18 to 48 h, and the sedation period is shaken once every 2 to 4 h; 2) After mixing the antigen saponin mixture with the ISA201VG adjuvant , Emulsification to obtain goat infectious pleuropneum sub-unit vaccine.

[0044]In the present invention, the goat infectious pleuropneumonia immunoprotein combination and saponin adjuvant are mixed, and the antigen saponin mixture is obtained. In the present invention, the preferred step of the following steps prior to the mixing of the goat pneumonia and saponin in the goat pneumonia in the goat, and the following steps: the following steps: A, B, C, Ding and Ding and Ding and Dinghe, a combination of ...

Embodiment 1

[0054]Preparation of meth protein

[0055]The MCCP metroid gene (GenBank: CP017125.1, M1601_02645, from 5 'ends No. 172-1524-bit nucleotide sequence, hypotheticalprotein), the codena optimization is synthesized to PET30A (+) restriction enzyme dug point NDEI and The protein shown in SEQ ID NO.1 is expressed between XHOI. The recombinant expression plasmid was obtained to transform Escherichia coli BL21 (DE3) to obtain recombinant bacteria. The recombinant bacteria were inoculated with LB liquid medium (containing 50 μg / ml kanamycin), 37 ° C 200 rpm was cultured to OD600It was 0.6, the final concentration was 0.05 mm IPTG, 16 ° C, 120 rpm induced 12h, and obtained recombinant expression of the bacterial liquid, 4 ° C, 12000 rpm centrifuge 30 min collected bacteria precipitation, pre-cooling Lysis buffer (50 mMNA)2HPO4, 0.3 mNaCl, 10 mmidazole, pH = 8.0) Heavy suspension precipitate, ultrasonic crushed 30 min (ultrasound 2S, interval 2S), 4 ° C, 7500 rpm centrifuge for 30 min, collecte...

Embodiment 2

[0065]Example 1 was immunized by the methyl protein, ethyl protein, propotein, butcobin, and penthenin, respectively, 3, and preparation of multi-cloned antibody, premium, preparation, metin, ethyl protein, propin, butter protein and The glutin is mixed with Furtol complete adjuvant and other volume, and the subcutaneous is more immunized, and each immunized approximately 800 ug recombinant protein; 2 weeks later, three weeks after 3 weeks, three weeks, three weeks, strengthen immunity Both are methyl protein, ethyl protein, propin, butrotein, and penthenin, each of the emulsified emulsions after mixing with Frequinth, each other, each immunized 600 μg of recombinant protein, last immunized day 7, intravenous blood Separate serum. Preparative metroids, ethyl protein, propin, butamopenoprotein polyclonal antibody 1: 80000 dilutions are also positive with corresponding recombinant protein Western, and the results show that the metroids, ethyl protein, propin, butcupin and The glutinin...

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Abstract

The invention provides a goat contagious pleuropneumonia subunit vaccine as well as a preparation method and application thereof, and belongs to the technical field of preparation of animal infectious disease vaccines, and a mycoplasma capricolum subsp.capripneumonia immune protein combination comprises mycoplasma capricolum subsp.capripneumonia immune proteins A, B, C, D and E; the mass ratio of the mycoplasma capricolum subsp.capripneumoniae immune proteins A to B to C to D to E is (0.5 to 1.5): (0.5 to 1.5): (0.5 to 1.5): (0.5 to 1.5): (0.5 to 1.5); and the goat contagious pleuropneumonia subunit vaccine comprises the goat mycoplasma goat pneumonia subspecies immune protein combination and an adjuvant. The subunit vaccine has the advantages of high safety, good immune effect, good stability and small side reaction.

Description

Technical field[0001]The present invention belongs to the field of animal infectious disease vaccine, and in particular, the present invention relates to a vaccine and preparation method thereof of goat infectious pleural pulmonary subunit.Background technique[0002]Goat infectious pleuropneumonia, CCPP is an important animal epidemic in the world's raising sheep industry, which is OIE statutory report infectious disease. The main clinical manifestation of goat infectious pleuropneumitis is fever, cough, difficulty breathing, and the cutting disease is mainly cellulose sex pleurulum pneumonia, pleural effusion and lung liver sample change. Goat infectious pleuropneumitis is a highly exposed to infectious disease. It has the characteristics of high morbidity and high mortality in the new disaster area, bringing serious threats and huge economic losses to raising sheep industry. Wild goats, Tibetan antelope and other animals can infect onset. Goat infectious pleuropneumitis is distribu...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/02A61K39/39A61P31/04
CPCA61K39/0241A61K39/39A61P31/04A61K2039/552A61K2039/55577A61K2039/55588A61K2039/55566
Inventor 储岳峰陈胜利郝华芳颜新敏马丽娜刘永生
Owner LANZHOU INST OF VETERINARY SCI CHINESE ACAD OF AGRI SCI
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