A probe composition for detecting drug-induced deafness gene and its application

Abstract

The present invention provides a probe composition for detecting drug-induced deafness gene and its application. The probe composition includes a first double-stranded detection probe for detecting the C1494T site, and a second double-stranded detection probe for detecting the A1555G site. Detection probe. The present invention uses the hybrid oligonucleotide of the site to be detected and its complementary oligonucleotide to form a double-stranded detection probe to detect the site to be detected, combined with a high-resolution melting curve, according to the melting curve of the double-stranded detection probe The change of the characteristic peak and the Tm value realizes the purpose of detecting different genotypes in one reaction system, the result is intuitive and clear, and the accuracy is high.

Description

technical field [0001] The invention belongs to the field of biotechnology and the field of gene detection technology, and relates to a probe composition for detecting drug-induced deafness genes and its application, in particular to a probe composition and reagents for detecting mutations of drug-induced deafness genes C1494T and A1555G Boxes, detection methods and their applications. Background technique [0002] At present, infectious diseases are one of the leading causes of death in the world, and more than 17 million people die from bacterial infections every year in the world. Aminoglycoside antibiotics, because of their good antibacterial activity and broad-spectrum antibacterial properties, are widely used to treat infections caused by Gram-negative and Gram-positive bacteria. However, it has been found in recent years that irrational use of aminoglycoside antibiotics can cause ototoxicity and nephrotoxicity. [0003] Carriers of mitochondrial DNA 12S rRNA gene C1...

AI Technical Summary

Problems solved by technology

However, these methods have certain limitations

Sequencing methods have high accuracy, predictability for unknown mutations, and high throughput of gene chips, but they all require specific instruments and reagents, and the cost is high, which is not conducive to popularization and application in underdeveloped areas; PCR-RFLP method requires PCR amplification first. Amplification, and then uncapping the amplified product may cause contamination and false positive results; the difference between the Tm values ​​of the wild type and the mutant type detected by the high-resolution melting curve method is small, which is easy to cause misjudgment and is difficult to detect. Achieve the effect of detecting two sites in the same reaction system

[0006] CN109234380A discloses a hereditary deafness-related gene detection kit and a specific primer set, although the kit can detect mutation sites of 33 hereditary deafness-related genes, including m.A1555G of the drug-induced deafness susceptibility gene 12S rRNA gene , m.C1494T locus, but it is developed based on a sequencing platform, and it needs to construct a library before sequencing and sequence comparison analysis. The operation is cumbersome, time-consuming, and costly, and it is easy to cause cross-contamination

[0007] CN105349624A discloses a hereditary deafness gene detection kit, and CN102787169A discloses a mixed solution, a kit and a detection system for detecting mutations of mitochondrial DNA A1555G and C1494T. The results are easily affected by factors such as sample concentration and purity, the accuracy is low, and the number of detection sites is limited by the detection channel, the operation is relatively cumbersome, and the results are not intuitive

[0008] The joint detection kit for deafness susceptibility genes disclosed in CN102864232A uses multiplex PCR combined with diversion hybridization technology to detect 13 mutation sites of four common deafness susceptibility genes (mtDNA, GJB2, GJB3 and SLC26A4) and their normal controls, However, after the PCR is completed, the method still needs to open the cover to take out the product for hybridization detection. The operation is relatively cumbersome and time-consuming, and the operation of opening the cover is easy to cause pollution.

[0009] Existing gene detection methods generally have the problems of cumbersome operation, long time-consuming, low throughput, easy pollution or high price. Therefore, a new method with simple and fast operation, high sensitivity and specificity, high throughput, and An economical detection method to achieve the purpose of simultaneously detecting the C1494T and A1555G mutations of 12S rRNA related to drug-induced deafness in the same reaction system

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