Application of brucellosis A19 molecular marking vaccine and immunological identification thereof
A technique for molecularly labeling vaccines and brucellosis, applied to bacterial antigen components, analytical materials, antibacterial drugs, etc., can solve the problem of difficult identification of immunized animals and clinically diseased animals
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Embodiment 1
[0027] The application of brucellosis A19 molecular marker vaccine and its immune protection:
[0028] Material:
[0029] Strains: A19-ΔVirB12 labeled vaccine was developed and preserved by the Veterinary Research Institute of Xinjiang Academy of Animal Sciences, Brucella 2308 was purchased from China Veterinary Drug Control Institute, and A19 vaccine was purchased from Xinjiang Tiankang Animal Husbandry Biotechnology Co., Ltd.;
[0030] Experimental animals: 25 Xinjiang native cattle aged 6-8 months, purchased from the live cattle trading market in Miquan District, Xinjiang, all tested negative for brucellosis;
[0031] Determination of the challenge dose of Brucella 2308 strain: 25 Xinjiang local cattle were randomly divided into 5 groups, 5 cattle in each group, among which, the 1-4 groups were challenged with Brucella 2308 standard strain, eyelid The inoculated challenge dose was 1×10 6 CFU, 1×10 7 CFU, 1×10 8 CFU, 1×10 9 CFU, the fifth group is the blank control grou...
Embodiment 2
[0042] Immunological identification of brucellosis A19 molecular marker vaccine:
[0043] Preparation of VirB12 antigen protein:
[0044] Strains and vectors: A19 molecularly labeled vaccine was developed by the Veterinary Research Institute of Xinjiang Academy of Animal Sciences; Brucella 2308 strains were purchased from the China Veterinary Drug Administration; the cloned pGEM-T vector was purchased from Promega as a commercial reagent; prokaryotic expression vectors pET-28a(+) is preserved by the Veterinary Research Institute of Xinjiang Academy of Animal Husbandry Science;
[0045] Reagents: Restriction enzymes were purchased from Fermentas; plasmid DNA extraction kit and DNA gel recovery kit were purchased from Promega; T4DNA ligase, DNA Marker, and DH5α were products of Beijing Dingguo Biotechnology Company. The enzyme-labeled secondary antibody was rabbit anti-bovine IgG horseradish peroxidase (Bethyl, USA); the His-trap column for protein purification was a product of...
Embodiment 3
[0054] iELISA identification of A19-ΔVirB12 vaccine antibody and field strain antibody:
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