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Preparation and applications for mycobacterium tuberculosis PUP protein overexpression strains

A tubercle bacillus, overexpression technology, applied in the field of genetic engineering and vaccine research, can solve the problem of lack of vaccines to prevent tuberculosis and other issues

Pending Publication Date: 2019-08-06
SHIHEZI UNIVERSITY
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  • Abstract
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  • Claims
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Problems solved by technology

The tuberculosis epidemic has not been controlled, and the root cause is the lack of effective vaccines to prevent tuberculosis

Method used

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  • Preparation and applications for mycobacterium tuberculosis PUP protein overexpression strains
  • Preparation and applications for mycobacterium tuberculosis PUP protein overexpression strains
  • Preparation and applications for mycobacterium tuberculosis PUP protein overexpression strains

Examples

Experimental program
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Effect test

Embodiment 1

[0028] Example 1 Preparation of Mycobacterium tuberculosis ubiquitin-like protein PUP overexpression strain

[0029] 1. Materials:

[0030] 1.1 Main materials:

[0031] Main materials: Bacillus Calmette-Guerin strain (referred to as BCG strain) is provided by the China Institute for the Control of Pharmaceutical and Biological Products and preserved in this laboratory; Escherichia coli E. coli DH5α was provided by our laboratory. The Escherichia coli-Mycobacterium tuberculosis shuttle expression vector pMV361 is preserved by our laboratory.

[0032] 1.2 Main reagents

[0033] 1) Restriction endonuclease EcoRI, American NEB Biological Company

[0034] 2) Restriction endonuclease Hind III, American NEB Biological Company

[0035] 3) T4 DNA Ligase American NEB Biological Company

[0036] 4) 2×pfu Master Mix Beijing Kangwei Century Company

[0037] 5) DNA Marker Beijing Tiangen Biochemical Technology Co., Ltd.

[0038] 6) Agarose gel recovery kit (spin column type) Beiji...

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Abstract

The invention discloses preparation and applications for mycobacterium tuberculosis PUP protein overexpression strains. BCG strains are cultivated, and the genome DNAs thereof are extracted. The genome DNAs of the BCG strains are taken as a template, and the PCR amplification of a mycobacterium tuberculosis Pup protein gene can be achieved; and sequencing and identification is performed. Escherichia coli-mycobacterium tuberculosis shuttle plasmid pMV361 is taken as a carrier, and recombinant shuttle plasmid pMV361-Pup : BCG is constructed and identified. An electroporation technology is utilized to transfer the constructed escherichia coli-mycobacterium tuberculosis shuttle plasmid pMV361 carrier (recombinant shuttle plasmid pMV361-Pup : BCG) into the BCG strains to construct and identifymycobacterium tuberculosis Pup protein overexpression strains which can be named as BCG : Pup strains. The strains can be used for studying the effects of ubiquitin-like proteasome system related protein PUP of mycobacterium tuberculosis on growth, drug resistance, permeability regulation, acid resistance and pathogenicity in hosts, and can also be used for the development and research of novel vaccines for intervening in and treating tuberculosis.

Description

technical field [0001] The invention relates to the fields of genetic engineering and vaccine research, and relates to the preparation and application of a bacterial strain overexpressing the PUP protein of Mycobacterium tuberculosis. Background technique [0002] Tuberculosis is a worldwide infectious disease caused by Mycobacterium Tuberculosis infection, and it is also the infectious disease with the highest mortality rate caused by a single bacterial infection. Tuberculosis is still the number one killer of infectious diseases in the world. New data released by the World Health Organization's "Global Tuberculosis Report 2017" shows that there were an estimated 10.4 million new cases worldwide in 2017, including 5.9 million (56%) males, 3.5 million (34%) females, and 1 million children. (34%). Six countries—India, Indonesia, China, Nigeria, Pakistan and South Africa—accounted for 60% of the total new cases. The root cause of tuberculosis epidemic failure is the lack of...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/74C12N1/21C12R1/32
CPCC12N15/74C07K14/35
Inventor 张万江赵正涌吴芳吴江东张杰董江涛徐芳柳小玲梁粟王洪洲赵彦恒邵萌
Owner SHIHEZI UNIVERSITY
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