Kit and method for detecting polymorphism of CYP3A4 gene

A gene polymorphism and kit technology, applied in the direction of biochemical equipment and methods, microbial measurement/inspection, etc., can solve the problems of CYP3A4 gene polymorphism, such as long time, long time, cumbersome steps, etc.

Active Publication Date: 2013-12-25
刘辉
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] In view of the problems in the prior art that the detection of CYP3A4 gene polymorphism takes a long time, the steps are cumbersome, and multiple PCRs are required, the purpose of the present invention is to provide a kit for detecting CYP3A4 gen

Method used

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  • Kit and method for detecting polymorphism of CYP3A4 gene
  • Kit and method for detecting polymorphism of CYP3A4 gene
  • Kit and method for detecting polymorphism of CYP3A4 gene

Examples

Experimental program
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Effect test

Embodiment 1

[0071] Embodiment 1. Preparation of kit

[0072] 1. Primer design for internal reference gene ACTB and CYP3A4 gene

[0073]According to the gene sequence, the ACTB gene sequence and the CYP3A4 gene sequence are derived from the National Center for Biotechnology Information Nucleic Acid Database (NCBI), wherein the ACTB gene ID is 60, and the reference sequence number is NG_007992.1, also refer to the SEQ ID NO in the sequence list of the present invention :35; the CYP3A4 gene ID is 1574, and the reference sequence number is NG_008421.1. Primer5.0 primer design software was used to design specific primers for the internal reference gene ACTB and the CYP3A4 gene, including: the long fragment of the 1st to 3rd exons of the CYP3A4 gene Amplification-specific upstream and downstream primers, long fragment amplification specific upstream and downstream primers of exons 4 to 7 of CYP3A4 gene, long fragment amplification specific upstream and downstream primers of exons 8 to 11 of CYP...

Embodiment 2

[0122] Example 2. Detection of CYP3A4 gene polymorphism with the kit prepared in Example 1

[0123] Take the results of random detection of peripheral blood samples from 30 subjects as an example.

[0124] The detection process of using the kit of the present invention to detect the CYP3A4 gene polymorphism in the population is as follows: first obtain the peripheral blood sample of the clinical subject, and quickly extract the genomic DNA; secondly, first prepare the PCR reaction solution of the internal reference gene ACTB, and then add the concentration of 2 ng / μl of internal reference gene ACTB sequence and 2 μl of internal positive control sequence were used for PCR amplification. After PCR, electrophoresis was performed in agarose DNA gel with a concentration of 2% to check the PCR amplification. If there are two bands of about 390bp and 800bp in the PCR amplification product of the internal reference gene ACTB, it indicates that the entire detection process is effective...

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Abstract

The invention relates to a kit and a method for detecting the polymorphism of a CYP3A4 gene and belongs to the technical field of gene sequencing. The kit comprises primers for detection, wherein the primers for the detection comprise at least one pair in long-fragment amplification specific forward and reverse primers for a first exon to a third exon of the CYP3A4 gene, long-fragment amplification specific forward and reverse primers for a fourth exon to a seventh exon of the CYP3A4 gene, long-fragment amplification specific forward and reverse primers for an eighth exon to an eleventh exon of the CYP3A4 gene and long-fragment amplification specific forward and reverse primers for a twelfth exon and a thirteenth exon of the CYP3A4 gene and at least one pair in forward and reverse sequencing primers for each exon of the CYP3A4 gene corresponding to long-fragment deoxyribose nucleic acid (DNA) of the first exon to the third exon, long-fragment DNA of the fourth exon to the seventh exon, long-fragment DNA of the eighth exon to the eleventh exon and long-fragment DNA of the twelfth exon and the thirteenth exon. According to the kit and the method, the detection time is short and the workload is low.

Description

technical field [0001] The invention relates to the technical field of gene sequencing, in particular to a kit and method for detecting CYP3A4 gene polymorphism. Background technique [0002] Cytochrome P450 (CYP), also known as mixed function oxidase, monooxygenase or drug metabolizing enzyme, is widely distributed in animals, plants and microorganisms. More than 20 isoenzymes have been found in mammalian tissues, which are divided into 4 families of CYP1-4. CYP3A-type enzymes account for 70% of the total CYP450 in the body, 30% of the CYP450 in the liver, and participate in the metabolic transformation of more than 60% of commonly used drugs in the body. At present, four CYP3A~ genes have been found in the human body, namely CYP3A4, CYF3A5, CYP3A7 and CYP3A43, and CYP3A4 is one of the most clinically important P450 enzymes, involved in the oxidative metabolism of more than 150 drugs in 38 categories, Accounting for about 50% of all drugs, it is the root cause of adverse ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 刘辉
Owner 刘辉
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