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Recombinant protein for diagnosing bovine tuberculosis and application thereof

A technology of recombinant protein and tuberculosis, applied in the field of recombinant protein, can solve the problems of cumbersome preparation of PPD, achieve the effect of maintaining natural activity, increasing the cost of biosafety control, and low cost

Active Publication Date: 2010-03-31
ZHAOQING DAHUANONG BIOLOGIC PHARMA
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The purpose of the present invention is to provide a highly specific and highly sensitive PPD replacement recombinant protein for the non-specificity of bovine PPD and the cumbersome preparation of PPD for the diagnosis of bovine tuberculosis

Method used

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  • Recombinant protein for diagnosing bovine tuberculosis and application thereof
  • Recombinant protein for diagnosing bovine tuberculosis and application thereof
  • Recombinant protein for diagnosing bovine tuberculosis and application thereof

Examples

Experimental program
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Embodiment 1

[0042] 1. Amplification of the target gene

[0043] The present invention adopts the technique of gene splicing by over lap extension, that is, the overlapping PCR method realizes the splicing of the target gene through two PCRs. A total of 4 primers were designed, P1 is the upstream primer of CFP10; the first half of P2 (the underlined part) is the linker sequence (Linker, encoding 15 hydrophobic amino acids), and the second half is the downstream primer of the ESAT6 gene; the first half of P3 (the underlined part) It is the base sequence (Linker) complementary to the underlined part of the primer P2, and the second half is the upstream primer of the ESAT6 gene; P4 is the downstream primer of the ESAT6 gene.

[0044] First, use primers P1 and P2 to amplify the CFP10 gene, use primers P3 and P4 to amplify the ESAT6 gene, then take the two gene products and mix them in equal volumes as a template, and use primers P1 and P4 to amplify the CFP10-ESAT6 gene.

[0045] The specific...

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Abstract

The invention discloses a recombinant protein for diagnosing bovine tuberculosis, which is obtained by a method comprising the following steps: extracting the genome DNA of mycobacterium tuberculosisvar. bovis as a template of a PCR reaction to amplify out CFP10 and ESAT6 genetic fragments; using a gene splicing technique to amplify out a target gene CFP10-ESAT6 fusion gene; cloning and transforming the target gene to obtain a recombinant fungus, and inducing and expressing the recombinant fungus according to a conventional method, and purifying an expression product to obtain the recombinantprotein. The recombinant protein and a diagnostic reagent provided by the invention are used for diagnosing the bovine tuberculosis to obtain the advantages of low cost, high speed, high sensitivityand specificity.

Description

technical field [0001] The invention relates to a recombinant protein, in particular to a recombinant protein for bovine tuberculosis diagnosis and its application. Background technique [0002] Bovine tuberculosis is a chronic wasting infectious disease of cattle mainly caused by Mycobacterium bovis. This disease can take place throughout the year, is the world epidemic, and in the severe epidemic area, the infection rate can reach 60%. More than a dozen countries in the world, including the United States, Canada, and Australia, have reported tuberculosis-free cattle. However, in recent years, due to the occurrence and prevalence of tuberculosis in wild animals and humans, the cattle herds in these countries with good tuberculosis control are also in constant decline. In quarantine and on high alert. Bovine tuberculosis not only leads to the decline of the productivity of the cattle industry itself, but more importantly, it poses a serious threat to human health. Humans ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K19/00C12P19/34C12N15/70C12P21/02G01N33/569C12R1/19C12R1/32
Inventor 陈瑞爱徐家华
Owner ZHAOQING DAHUANONG BIOLOGIC PHARMA
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