Method and kit for detecting polymorphism of ADPRT gene

A technology of gene polymorphism and detection method, which is applied in the field of detection method and the kit of the method, can solve the problems of unsuitable small sample mutation detection, high detection cost, expensive equipment, etc., and achieve high specificity and good repeatability , the effect of saving samples

Inactive Publication Date: 2009-11-04
夏昭林 +2
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Problems solved by technology

[0006] In recent years, people have developed many methods for detecting gene polymorphism, the more common ones are single-strand conformation polymorphism (SSCP), direct sequencing technology, denaturation-high pressure liquid chromatography (DHPLC), dynamic allele-specific Hybridization techno

Method used

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Embodiment 1

[0072] Using the genomic DNA extracted above as a template, design gene sequence forward and reverse primers in the region where the ADPRT gene locus Val762Ala (rs1136410) is located. The forward primers are: 5'-TTTTGCTCCTCCAGGCCAAC*G-3'(before * The base is the introduced mismatched base to create a restriction site); the reverse primer is: 5'-CCTGACCCTGTTACCTTAATGTCAGTTTT-3', a restriction site is introduced into the forward primer through a single base mismatch, And carry out specific PCR amplification to the template genomic DNA; The specific PCR amplification steps are as follows:

[0073] For the sample genomic DNA, use the above-mentioned forward and reverse primers to carry out PCR amplification according to the following reaction system and conditions: 25μl reaction system, including 1U Taq DNA polymerase, 10×Buffer2.5μl, 25mol / L MgCl22μl, 2.5mmol / L dNTPs 2.5 μl, 50ng human genome DNA, 10 μmol / L upstream and downstream primers 1 μl each, and 25 μl made up with sterili...

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Abstract

The invention discloses a method for detecting the polymorphism of an ADPRT gene, which is characterized by comprising the following steps: (a) human genomic DNA is used as a template, and a positive primer and a reverse primer of a gene sequence are designed in a region of an ADPRT gene locus; a single basic group is mismatched on the positive primer so as to introduce an enzyme cutting site; and the template genomic DNA is processed by specificity polymerase chain reaction (PCR) amplification; (b) a restriction enzyme is utilized to carry out enzyme cutting on the genomic DNA; and (c) segment size separation is carried out on the ADPRT gene according to agarose gel electrophoresis so as to determine the gene polymorphism of the ADPRT gene. The invention also discloses a kit for detecting the ADPRT gene polymorphism, which comprises a gene polymorphism primer for amplification, dNTPs, DNA polymerase for the PCR reaction, a buffer solution of the DNA polymerase, a restriction enzyme Bsh1236I for restriction fragment length polymorphism (RFLP) and a corresponding buffer solution. The method has simple operation, low cost, intuitional judging result, quick reaction and convenient popularization and application, and the kit is convenient to use.

Description

technical field [0001] The invention relates to a gene detection method, in particular to a detection method for detecting ADPRT gene polymorphism and a kit for the method. Background technique [0002] The occurrence and evolution of tumors is a multi-stage and multi-step process of interaction between environmental factors and genetic factors. From a macroscopic point of view, this process reflects the interaction between the environment and the host, especially the genetic factors of the host; while from the microscopic point of view, the occurrence of tumors essentially reflects the influence and effect of carcinogenic factors on the genetic material of cells. . Studies have shown that among people living in the same environment, some suffer from cancer, while others are safe. Even in some areas with high tumor incidence, the actual incidence of tumor is only about 0.1-0.2%. Over the years, in the study of tumor etiology, a large amount of data has been accumulated, in...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 夏昭林缪文彬张忠彬
Owner 夏昭林
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