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Reagent kit for detecting pathogenic mycobacterium tuberculosis through uniting multiplex PCR, nested-PCR and touch-down PCR

A Mycobacterium tuberculosis, pathogenic technology, applied in the field of biomedicine, can solve problems such as inability to meet accurate detection, genotype differences, etc.

Active Publication Date: 2019-11-12
天津市泌尿外科研究所
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, only relying on a conservative gene or protein detection method can no longer meet the needs of accurate detection.
Moreover, wild-type combined mycobacteria and BCG (BCG) for routine immunization also showed significant differences in genotype

Method used

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  • Reagent kit for detecting pathogenic mycobacterium tuberculosis through uniting multiplex PCR, nested-PCR and touch-down PCR
  • Reagent kit for detecting pathogenic mycobacterium tuberculosis through uniting multiplex PCR, nested-PCR and touch-down PCR
  • Reagent kit for detecting pathogenic mycobacterium tuberculosis through uniting multiplex PCR, nested-PCR and touch-down PCR

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Experimental program
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Effect test

Embodiment Construction

[0033] For verifying technical effect of the present invention, carry out following experiment:

[0034] 1 Materials and methods

[0035] 1.1 Materials

[0036] 1.1.1 Samples: Mycobacterium tuberculosis samples from clinically confirmed tuberculosis patients, standard wild-type pathogenic Mycobacterium tuberculosis (H37RV), BCG (Danish strain), BCG (Pasteur strain), Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa, Streptococcus pneumoniae

[0037] 1.1.2 Main reagents 1; DNA extraction kit, (provided by Tianjin New Platform Biotechnology Co., Ltd.) 2; MasterMix, 3; ultrapure water, 4; primers, 5; TAE, 6; EB

[0038] 1.2 Method

[0039] 1.2.1 Primer design and synthesis Referring to the standard Mycobacterium tuberculosis (H37RV) genome published by GenBank as a template, and according to the design principles of multiplex PCR and nested PCR primers, primers for IS1081, IS6110 and MPB64 were designed by Tianjin Jinweizhi Company Synthesis. See Table 1.

[0...

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Abstract

The invention relates to a reagent kit for detecting pathogenic mycobacterium tuberculosis through uniting multiplex PCR, nested-PCR and touch-down PCR. The reagent kit adopts a technique uniting themultiplex PCR, the nested-PCR and the touch-down PCR, and through 3 groups of primers of IS6110, IS1081 and MPB64, 3 molecule markers of IS6110, IS1081 and MPB64 are detected in a multiplex, efficientand specific manner, so that the pathogenic mycobacterium tuberculosis can be rapidly diagnosed, and determination can be more precise. The reagent kit can detect conservative molecule markers including IS6110 and IS1081 of the pathogenic mycobacterium tuberculosis, besides, can detect MPB64 molecule markers in a wild strain of the pathogenic mycobacterium tuberculosis, and further has accurate judgment significance in the pathogenic mycobacterium tuberculosis, and the detection right rate of the pathogenic mycobacterium tuberculosis can be substantially increased.

Description

technical field [0001] The invention relates to a kit for detecting pathogenic Mycobacterium tuberculosis by combining multiplex PCR nested PCR and touch down PCR, which belongs to the technical field of biomedicine. Background technique [0002] Tuberculosis is a serious zoonotic infectious disease and one of the top ten causes of death worldwide. According to the World Health Organization, in 2017, 10 million people worldwide suffered from tuberculosis, and 1.6 million people died of tuberculosis. Tuberculosis poses a serious threat to both human health and livestock production. [0003] Tuberculosis is mainly pulmonary tuberculosis, caused by Mycobacterium tuberculosis in the lungs. The traditional primary diagnosis of pulmonary tuberculosis relies on sputum smear, acid-fast staining and PPD test, the sensitivity and specificity are not high. Due to the slow growth of Mycobacterium tuberculosis, sputum cultures took 8 weeks, delaying the treatment of tuberculosis. In ...

Claims

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Application Information

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IPC IPC(8): C12Q1/6848C12Q1/689C12Q1/04C12N15/11
CPCC12Q1/6848C12Q1/689C12Q2600/16C12Q2531/113C12Q2537/143C12Q2549/119Y02A50/30
Inventor 牛远杰国林沛陈家童郭明日尚芝群唐慧琴
Owner 天津市泌尿外科研究所
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