Schistosoma japonicum recombinant multi-epitope antigens, method for expressing and purifying same and application thereof

A schistosomiasis, multi-epitope technology, applied in the direction of biochemical equipment and methods, applications, recombinant DNA technology, etc., can solve the problems of specificity, unstable repeatability, low detection rate, and long time spent

Inactive Publication Date: 2010-01-13
SHANGHAI VETERINARY RES INST CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The pathogenic diagnosis method of cattle, sheep and other large livestock is currently widely used by nylon sieve elutriation to collect eggs combined with fecal miracidium hatching. This method has reliable results, but the detection rate i

Method used

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  • Schistosoma japonicum recombinant multi-epitope antigens, method for expressing and purifying same and application thereof
  • Schistosoma japonicum recombinant multi-epitope antigens, method for expressing and purifying same and application thereof
  • Schistosoma japonicum recombinant multi-epitope antigens, method for expressing and purifying same and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0086] Example 1. Construction of Schistosoma japonicum multi-epitope recombinant nucleic acid vaccine and immune protection experiment

[0087] 1. Materials

[0088] 1.1 Plasmids and strains: eukaryotic expression vector pCMV-script, Escherichia coli DH5α, etc. were purchased from Shanghai Shenneng Biotechnology Company.

[0089] 1.2 Experimental animals: male Kunming mice, weighing 25 g, purchased from Shanghai Experimental Animal Center.

[0090] 1.3 Cercariae of Schistosoma japonicum mainland China strain: Provided by the Oncomelania Room of Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences.

[0091] 1.5 Main reagents: ampicillin, kanamycin, Agarose, Tryptone, yeast extract, NaCl, etc. were purchased from Shanghai Sangon Bioengineering Co., Ltd.; DNA Marker DL2000, dNTP Mixture, MgCl 2 、Taq TM DNA Ployrose, restriction enzymes EcoRI, HindIII, BamHI, XbaI, T 4 DNA ligase ligase, etc. are products of Dalian Bao Biotechnology Co., Ltd. DNA ...

Embodiment 2

[0157] Example 2, Expression, Purification and Immunoprophylaxis of Schistosoma japonicum Multi-epitope Recombinant Antigen

[0158] 1. Materials

[0159] 1.1 Plasmids and strains: prokaryotic expression vector pGEX-2T, Escherichia coli DH5a, BL21, etc. were purchased from Shanghai Shenneng Biotechnology Company.

[0160] 1.2 Experimental animals: BALB / c mice: male, 6 weeks old, purchased from Shanghai Experimental Animal Center.

[0161] 1 and 3 The cercariae of Schistosoma japonicum were provided by the snail laboratory of Shanghai Institute of Livestock Parasitic Diseases, Chinese Academy of Agricultural Sciences.

[0162]1.4 Enzymes and other related reagents: Ampicillin, IPTG, X-gal, glutathione, urea, etc. were purchased from Shanghai Sangon Bioengineering Company. DNA gel recovery kit, plasmid extraction kit, standard protein marker, etc. were purchased from Tianwei Times Bio Company, and protein refolding kit was purchased from Pufei Bio Company. Taq DNA polymerase,...

Embodiment 3

[0193] Example 3, Recombinant multi-epitope antigen is used for the research experiment of diagnosing bovine schistosomiasis

[0194] 1. Materials

[0195] 1.1 Antigens: recombinant multi-epitope antigens pGEX-BSjGCP-BSj23 and pGEX-BSjGCP-BSj23-BSj28 were prepared according to the aforementioned method. Store at -20°C for later use.

[0196]1, 2 Serum: Buffalo schistosomiasis-positive serum was collected from grazing buffaloes in the schistosomiasis-endemic areas of Yunnan Province. Before collection, the local veterinarians checked the feces of target buffaloes by the hatching method of fecal miracidia according to the national standard method to diagnose schistosomiasis in buffaloes, and the serum of buffaloes with positive feces test was regarded as the positive sera of schistosomiasis. Serum from healthy bovines in Henan, a schistosomiasis-free area, was isolated and stored at -20°C for future use. Take 50 parts of healthy buffalo serum and mix them to make buffalo stan...

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Abstract

The invention discloses gene orders of schistosoma japonicum recombinant multi-epitope antigens BSjGCP-BSj23 and BSjGCP-BSj23-BSj28, a method for expressing and purifying the same, and application thereof in preparing schistosomiasis japonica immunity prevention vaccines and diagnostic reagents. Recombinant multi-epitope nucleic acid vaccines pCMV-BSjGCP-BSj23 and pCMV-BSjGCP-BSj23-BSj28 obtain 14.76 percent and 64.95 percent of worm reduction rates respectively in Kunming mice. The recombinant multi-epitope antigens pGEX-BSjGCP-BSj23 and pGEX-BSjGCP-BSj23-BSj28 obtain 15.7 percent and 57.99 percent of worm reduction rates in immunizing BalB/c mice, and obtain 91.0 percent and 89.9 percent of sensitivities as well as 97.8 percent and 93.4 percent of specificities respectively as diagnostic antigens.

Description

technical field [0001] The invention relates to biological genes, in particular to recombinant multi-epitope antigen of schistosomiasis japonicum and its expression, purification method and application in preparation of immune prevention vaccine and diagnostic reagent of schistosomiasis japonicum. Background technique [0002] Schistosomiasis japonicum is an important zoonotic parasitic disease. Because the intermediate host snails are difficult to eradicate and the repeated infection of schistosomiasis is serious, drug therapy alone cannot control the prevalence of schistosomiasis and eventually eliminate schistosomiasis. Therefore, research on anti-schistosomiasis vaccines has become one of the focuses of schistosomiasis research. Up to now, hundreds of schistosomiasis antigen genes have been cloned and identified, no less than 30 schistosomiasis gene recombinant antigen vaccines and nucleic acid vaccines have been used in animal immune protection tests, and some genetical...

Claims

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Application Information

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IPC IPC(8): C07K14/435C12N15/12C12N15/70A61K48/00A61P33/12G01N33/68
Inventor 林矫矫傅志强章登吉周伟芳陆珂石耀军刘金明李浩朱传刚
Owner SHANGHAI VETERINARY RES INST CHINESE ACAD OF AGRI SCI
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