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ALDH2 gene polymorphism detection kit based on self-quenched probe melting curve and ALDH2 gene polymorphism detection method based on self-quenched probe melting curve

A gene polymorphism and melting curve technology, which is applied in the field of ALDH2 gene polymorphism detection kits, can solve the problems of high price, low precision, unsuitable detection of single gene SNP loci, etc. Inexpensive effect

Inactive Publication Date: 2017-12-08
MENGCHAO HEPATOBILIARY HOSPITAL OF FUJIAN MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0021] Compared with traditional instrument detection methods, it has the characteristics of high throughput, miniaturization, automation, and anti-pollution. It is only suitable for whole-genome SNP scanning and is not suitable for SNP site detection of a single gene. It has low precision and is expensive.

Method used

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  • ALDH2 gene polymorphism detection kit based on self-quenched probe melting curve and ALDH2 gene polymorphism detection method based on self-quenched probe melting curve
  • ALDH2 gene polymorphism detection kit based on self-quenched probe melting curve and ALDH2 gene polymorphism detection method based on self-quenched probe melting curve
  • ALDH2 gene polymorphism detection kit based on self-quenched probe melting curve and ALDH2 gene polymorphism detection method based on self-quenched probe melting curve

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Embodiment 1

[0056] The specific sequence information of primers and probes is as follows:

[0057] Upstream primer F: 5'-TGGTGGCTACAAGATGTCG-3';

[0058] Downstream primer R: 5'-GCAGACCCTCAAGCCCCAAC-3';

[0059] Fluorescently labeled probe P:

[0060] 5’-FAM-T*G*C*AGGCATACACTGAAGTGAAAACTGTG-BHQ1-3’; where the bases marked with * are thio-modified.

[0061] Amplification reaction reagent PCR Mix, composed of: 1×PCR bufferA ((670mM Tris-HCl, 67uM EDTA, 166mM (NH 4 ) 2 SO 4 , 0.85mg / ml BSA)), 5mM MgCl 2 solution, 0.2mM dNTP solution, 0.02uM F, 0.4uM R, 0.3uM P.

[0062] The method of the kit for detecting ALDH2 gene polymorphism based on the self-quenching probe melting curve method comprises the steps:

[0063] S1: Sample DNA extraction

[0064] Take 10uL saliva sample and add it to 90uL DNA extraction solution (DNA-EZ Reagents V All-DNA-Out, Sangon Bioengineering (Shanghai) Co., Ltd.) Do not exceed 1 / 10 of the amount of DNA extraction solution), heat in a constant temperature wate...

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Abstract

The invention discloses an ALDH2 gene c.1510 site polymorphism detection kit and an ALDH2 gene polymorphism detection method based on self-quenched probe melting curve. The invention designs primers and a probe specifically for an ALDH2 gene, namely an upstream primer F:5'-TGGTGGCTACAAGATGTCG-3', a downstream primer R:5'-GCAGACCCTCAAGCCCCAAC-3', and a fluorescent marking probe P:5'-FAM-T*G*C*AGGCATACACTGAAGTGAAAACTGTG-BHQ1-3'. The kit disclosed by the invention has the advantages of being high in sensitivity, good in specificity, rapid in reaction and low in cost and is applicable to large-scale clinical application, rapid, effective and accurate parting qualitative detection on ALDH2 is achieved, and reference is provided for in-time treatment on angina with nitroglycerin, alcohol drinking instruction and related cancer occurrence.

Description

(1) Technical field [0001] The invention relates to an ALDH2 gene polymorphism detection kit and detection method based on the melting curve of a self-quenching probe. (2) Background technology [0002] Human acetaldehyde dehydrogenase 2 (ALDH2) is a key enzyme in alcohol metabolism, and its coding gene ALDH2 has a high degree of polymorphism. ALDH2 gene *2 polymorphism (NM_000690.3:c.1510G>A, p.Glu504Lys) can lead to a significant decrease in the enzyme activity of acetaldehyde dehydrogenase, and the metabolism of acetaldehyde into acetic acid is slower, resulting in the accumulation of acetaldehyde in the body after drinking. accumulation. When acetaldehyde in the blood reaches a certain concentration, there will be discomforts such as flushing, headache, tachycardia, and dyspnea, which are manifested as intolerance to alcohol, which is called flushing reaction. Especially the concentration of acetaldehyde in the blood of ALDH2*2 / ALDH2*2 genotype (homozygous mutant ty...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
CPCC12Q1/6858C12Q2531/113C12Q2527/107
Inventor 刘小龙刘景丰许海坡蔡志雄曾永毅董秀清陈耕
Owner MENGCHAO HEPATOBILIARY HOSPITAL OF FUJIAN MEDICAL UNIV