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A kit and method for identifying mycobacterial strains

A technology of mycobacteria and kit, applied in the field of molecular biology, can solve problems such as inability to identify

Active Publication Date: 2018-10-16
THE 309TH HOSPITAL OF CHINESE PEOPLES LIBERATION ARMY +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

After experiments, the inventors found that the method disclosed in this patent document only uses common double-labeled probes, and the difference in Tm values ​​between some bacterial species is small, and it is only applicable to the identification of a single bacterial species isolated and cultivated, while for two or the presence of multiple bacteria at the same time, it is impossible to identify

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  • A kit and method for identifying mycobacterial strains
  • A kit and method for identifying mycobacterial strains
  • A kit and method for identifying mycobacterial strains

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Experimental program
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Effect test

Embodiment 1

[0068] A method for identifying mycobacterial strains, the steps are as follows:

[0069] 1. Design and synthesis of primers and probes

[0070] The inventors compared and analyzed the 16S rRNA gene sequences of 12 common mycobacteria, and found the genus-specific and species-specific variable regions corresponding to the 12 kinds of mycobacteria: the first variable region (No. 1 region) is located between the 55-251 bases of the sequence X55588.1 in the NCBI database, the second variable region (the second region) is located between the 366-539 bases, and the third variable region (the third Region) is located between the 1110-1314th bases, although three specific probes are disclosed in the patent CN102634575B, these three specific probes are only ordinary double-labeled probes, and the difference in Tm value between some bacterial species is small , which is suitable for the identification of a single bacterial species in isolation and culture, but cannot be identified for...

Embodiment 2

[0120] 1. Detection of compound strains

[0121] In recent years, in skin soft tissue infection, skeletal system infection and 50% AIDS (acquired immunodeficiency syndrome) autopsy patients, it is found that avian-intracellular complex infection, according to the characteristics of the probe of the present invention, two nucleotide sequence differences The Tm value of the melting spectrum produced by the dots is quite different, so for the infection samples of the two mycobacterial complexes, double peaks can appear in the melting spectrum of the same tube reaction, that is, the melting peaks corresponding to the two strains.

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Abstract

The invention relates to a kit and a method for identifying mycobacterium strains. The method comprises the following steps: (1) design and synthesis of a primer and a probe; (2) PCR reaction component; (3) PCR amplification; and (4) melting curve analysis. The kit and the method disclosed by the invention, on the basis of fluorescent PCR and melting curve analysis technical platform, can detect 12 mycobacterium strains which are common in the clinical field through the innovative application of the novel double-labeled and double-specific probe which has nucleic acid modification, so that the Tm value difference among the various strains is more than or equal to 4 DEG C within at least one detection channel; and the kit and the method is suitable for accurately and effectively identifying a composite strain that two or more strains simultaneously exist.

Description

technical field [0001] The invention relates to a kit and a method for identifying mycobacterium strains, belonging to the technical field of molecular biology. Background technique [0002] Polymerase chain reaction (PCR) technology has become a routine technology in laboratories after several decades of development since it came out in 1985. It is an indispensable means in the research of modern molecular biology, and it is an extremely sensitive amplification system. Compared with traditional clinical diagnostic methods, nucleic acid diagnosis is a diagnostic technology at the molecular level, which can make up for some shortcomings of traditional clinical diagnostic methods. Therefore, nucleic acid diagnostic technology represented by PCR technology has been increasingly widely used in clinical diagnosis. [0003] Compared with the traditional PCR detection mode, in recent years, the PCR amplification and detection in the traditional PCR detection mode have been combin...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/689C12Q1/686C12Q1/04
CPCC12Q1/686C12Q1/689C12Q2527/107C12Q2537/143C12Q2525/117Y02A50/30
Inventor 吴雪琼张俊仙汪君黄利维孙伟民
Owner THE 309TH HOSPITAL OF CHINESE PEOPLES LIBERATION ARMY