Multiplex PCR Kit for Tuberculosis and NTM Differentiation
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Solution Overview
Problem
Current methods for detecting Mycobacterium tuberculosis and nontuberculous mycobacteria are inaccurate due to shared nucleotide sequences, leading to misidentification and the need for a prompt and selective detection method that can differentiate between the two.
Innovation Solution
A method using specific primers and probes for the IS6110 gene of Mycobacterium tuberculosis and the 16S rRNA gene of nontuberculous mycobacteria, combined with multiplex real-time PCR, to selectively amplify and detect these pathogens in samples.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Adaptability or versatility
If nucleotide sequences that are unique to nontuberculous mycobacteria as well as sequences present in Mycobacterium tuberculosis are used in test reagents, then detection coverage is improved, but detection accuracy deteriorates due to cross-reactivity and misidentification
Solution Approach 1:
The invention divides the detection target into two distinct segments: Mycobacterium tuberculosis-specific sequences (IS6110 gene) and nontuberculous mycobacteria-specific sequences (16S rRNA gene). By using separate primer sets for each segment, the method achieves specific detection of each pathogen type without cross-reactivity, resolving the contradiction between detection coverage and accuracy
Solution Approach 2:
The invention applies different detection strategies tailored to each pathogen type: IS6110 gene detection for M. tuberculosis and 16S rRNA gene detection for nontuberculous mycobacteria. This localized approach allows optimization of detection conditions for each specific target, improving both coverage and precision simultaneously
2Measurement precision
If conventional culture and identification tests are used, then comprehensive identification is achieved, but detection time increases to 2-4 weeks
Solution Approach 1:
The invention replaces the mechanical culture-based identification system with a molecular biology-based PCR system. By using polymerase chain reaction to amplify and detect specific gene sequences (IS6110 for M. tuberculosis and 16S rRNA for nontuberculous mycobacteria), the method achieves rapid identification within hours rather than weeks, while maintaining or improving accuracy
Solution Approach 2:
The invention performs preliminary amplification of target gene sequences before detection. By pre-designing specific primers for IS6110 and 16S rRNA genes, the method enables direct and rapid identification from clinical samples without requiring extended culture periods, significantly reducing detection time while preserving identification accuracy
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
Enables efficient and accurate detection of Mycobacterium tuberculosis and nontuberculous mycobacteria, reducing misidentification and allowing for appropriate therapeutic selection.
Implementation Method 1
amplifying a target nucleotide sequence in the sample using: (i) a Mycobacterium tuberculosis detection set comprising a primer pair including a primer of SEQ ID NO: 1 and a primer of SEQ ID NO: 2, and at least one probe selected from the group consisting of SEQ ID NO: 3 and SEQ ID NO: 4
Implementation Method 2
amplifying a target nucleotide sequence in the sample using: (ii) a nontuberculous mycobacteria detection set comprising a primer pair including at least one primer selected from the group consisting of SEQ ID NO: 5, SEQ ID NO: 6, and SEQ ID NO: 7 and a primer of SEQ ID NO: 8, and at least one probe selected from the group consisting of SEQ ID NO: 9 and SEQ ID NO: 10
Data Source
AI summary
The present invention relates to a method for specifically detecting Mycobacterium tuberculosis and nontuberculous mycobacteria by simultaneously amplifying and analyzing target genes using various primers and probes, and a kit using same. The method of the present invention is capable of selectively detecting Mycobacterium tuberculosis and nontuberculous mycobacteria with very high efficiency through a multiplex real-time polymerase chain reaction (PCR) using probes and primers specific to target genes (particularly, IS6110, 16S rRNA and β-actin). Also, the kit of the present invention is capable of conveniently and efficiently detecting the target genes in a sample through a multiplex real-time PCR. Therefore, the method and the kit of the present invention are capable of selectively detecting with ease whether or not there is an infection with Mycobacterium tuberculosis or nontuberculous mycobacteria in a sample, and can be more accurately applied to the treatment of diseases on the basis thereof.