Hypertension gene polymorphism fluorescent PCR (polymerase chain reaction) solubility curve detecting kit and application thereof

Abstract

The invention relates to a hypertension gene polymorphism fluorescent PCR solubility curve detecting kit and application thereof, and a hypertension medication gene multiple-PCR solubility curve detecting method. The hypertension gene polymorphism fluorescent PCR solubility curve detecting kit mainly comprises a primer combination, a probe combination, and PCR reaction agents such thermally activated polymerases, PCR buffer solution and dNTPs (deoxy-ribonucleoside triphosphates) and involves polymorphism detection of 7 hypertension-related genes including CYP2D6*10, CYP2C9*3, ADRB1(1165G)C), AGTR1(1166A)C), CYP3A5*3, NPPA(T2338C) and ACE(I / D); the three reaction agents are applied to perform PCR amplification and solubility analysis to determine the related polymorphism of the 7 genes to guide clinical customized application of 5 majors types of hypertension drugs. The hypertension gene polymorphism fluorescent PCR solubility curve detecting kit can easily complete detection through 3reaction tubes, and meanwhile, save a nucleic acid extraction process, simplify operation steps and achieve a simple interpretation method.

Description

(1) Technical field [0001] The invention relates to a fluorescent PCR melting curve detection kit for gene polymorphism of hypertension, an application thereof, and a multiplex PCR melting curve detection method for detecting hypertension medication genes. (2) Background technology [0002] As a chronic non-communicable disease, hypertension is also a chronic disease with high prevalence, high disability rate and heavy disease burden in my country. According to data released by the National Health and Family Planning Commission in 2016, the prevalence of hypertension among adults aged 18 and over in my country is 25.2%. Although the awareness rate, treatment rate, and control rate of hypertension in the Chinese population have improved in recent years, they are still at a relatively low level. The global burden of disease study shows that the disability-adjusted life-year (DALY) of the Chinese population due to hypertension is as high as 37.94 million person-years, accounti...

AI Technical Summary

Problems solved by technology

[0008] At present, there are many technologies for gene polymorphism detection, the most commonly used are polymerase chain reaction-restriction fragment length polymorphism analysis (PCR-RFLP method), sequence-specific PCR, first-generation sequencing, next-generation sequencing and other technologies However, these technologies have their own application defects, some are cumbersome to operate, the detection cycle is long, and it is impossible to perform high-throughput multi-site simultaneous detection, which makes the current clinical application of polymorphism detection technology still unsatisfactory, and has not been able to meet clinical requirements. Testing needs

[0009] Multiplex fluorescent PCR combined with high-resolution probe method melting curve method is a new type of SNP analysis technology, which is improved on the basis of HRM (high-resolution melting curve) technology, and the saturated dye in HRM is replaced by a probe. The position of the probe design is on the SNP site, which makes the detection result more direct and specific, and the biggest advantage is that it can be used for typing detection. One probe can distinguish all polymorphisms of a SNP site, and combine multiple Color fluorescence detection greatly improves the detection throughput. However, one fluorescence channel of the traditional fluorescent PCR method can only detect one genotype. If the probe method and the melting curve method are used, one fluorescence channel can detect three genotypes or even more. The throughput is magnified by 3 times, and what needs to be explained is that the conventional fluorescent PCR method faces interference between subtypes, so most of the methods that use Ct value difference in interpretation make the detection results unstable

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