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A kind of recombinant BCG strain overexpressing Mycobacterium tuberculosis rv3586 and its application

A technology of Mycobacterium tuberculosis and recombinant BCG, applied in the field of tuberculosis vaccine

Active Publication Date: 2019-07-26
FOURTH MILITARY MEDICAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, rBCG that can replace traditional BCG has not yet been obtained, indicating that it is still necessary to make full use of existing research results, extensively screen new mechanism of action antigens, or improve vaccine construction methods or immunization strategies, and more effective rBCG and its immunization methods have been obtained

Method used

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  • A kind of recombinant BCG strain overexpressing Mycobacterium tuberculosis rv3586 and its application
  • A kind of recombinant BCG strain overexpressing Mycobacterium tuberculosis rv3586 and its application
  • A kind of recombinant BCG strain overexpressing Mycobacterium tuberculosis rv3586 and its application

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Experimental program
Comparison scheme
Effect test

Embodiment Construction

[0044] Construction of Rv3586 Escherichia coli-Mycobacterium Shuttle Expression Vector

[0045] Primers were designed according to the genome Rv3586 gene sequence of the Mtb H37Rv strain:

[0046] P1: 5'-tttaagcttatgcacgctgtgactcgtccgacc-3' HindIII site

[0047] P2: 5'-gcgaagcttaaggcggataattattgatcgc-3' HindIII site

[0048] Primers were synthesized by Shanghai Sangon Biotechnology Co., Ltd. Use the genomic DNA of Mtb H37Rv strain as a template to amplify the target gene by PCR. Reaction parameters: pre-denaturation at 95°C for 5 minutes, denaturation at 95°C for 30 seconds, renaturation at 57°C for 1 minute, extension at 72°C for 100 seconds, 35 cycles, and extension at 72°C for 5 minutes after the last cycle . 1% agarose gel electrophoresis analysis showed that a 1 100bp target band was amplified ( figure 1 ). The target fragment was recovered by the gel recovery kit, digested with HindIII, and the digested product was recovered by 1% agarose gel electrophoresis. T4 DN...

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Abstract

The invention relates to a recombinant BCG strain overexpressing Mycobacterium tuberculosis Rv3586 and application thereof. The plasmid containing the gene encoding Mycobacterium tuberculosis Rv3586 was transformed into BCG, and the strain obtained by screening was called Bacillus calmette-Guerin rBCG-DisA, or rBCG-DisA for short, and the preservation number was CCTCC M 2016335. Recombinant BCG-DisA overexpressing Rv3586 has the dual advantages of both target antigen and BCG. Immunization alone or in combination with Mycobacterium tuberculosis Ag85B-ESAT6 subunit vaccine can induce stronger immune responses in normal and infected mice and improve The protection of the body against mycobacteria has a good application prospect.

Description

technical field [0001] The invention belongs to the field of tuberculosis vaccines. The invention relates to a recombinant BCG strain overexpressing Mycobacterium tuberculosis Rv3586. The present invention also relates to the application of the recombinant BCG strain alone or in combination with the subunit vaccine Ag85B-ESAT6 (AE) in tuberculosis preventive and therapeutic vaccines and preparations. Background technique [0002] Tuberculosis Epidemic Status [0003] Tuberculosis (TB) is still the most serious infectious disease worldwide. Mycobacterium tuberculosis (Mtb) is the pathogen of TB, and about 1 / 3 of the world's population has been infected with Mtb. According to the 2014 WHO report, there were approximately 9.6 million new TB cases worldwide. my country is one of the 22 countries with a high TB ​​burden in the world. In 2014, the number of new TB patients in my country was 930,000, ranking third in the world after India and Indonesia. The incidence of TB is ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N1/21C12N15/74C12N15/60A61K39/04A61P31/06C12R1/32
CPCA61K39/04A61K2039/523C12N9/88C12Y406/01001C12N1/205C12R2001/32
Inventor 柏银兰徐志凯王立飞康健路延之宁唤唤吴俊杰宋世杰姚庆港
Owner FOURTH MILITARY MEDICAL UNIVERSITY
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