Method and kit for detecting CYP2C19 gene polymorphism

A CYP2C19, gene polymorphism technology, applied in the field of biotechnology and detection, can solve the problems of expensive probes, low throughput and high throughput

Pending Publication Date: 2019-03-01
DAAN GENE CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0008] (3) PCR-gene chip method, this method has high throughput and can detect multiple SNP sites to be tested at the same time, but the cost is high and special instruments and equipment are required
[0009] (4) Real-time fluorescent PCR method has high throughput, simple operation, and easy popularization of instruments and equipment, but the probes are more expensive
[0010] (5) PCR-restriction fragment length polymorphism method does not require special equipment, low cost, simple experimental process, strong operability, but low throughput, only suitable for partial SNP typing
[0011] (6) ARMS-PCR method, which can be used to detect various types of SNPs, has the advantage of high sensitivity, and is especially suitable for detecting somatic mutations in tumor tissues; the disadvantage is that false positives may occur

Method used

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  • Method and kit for detecting CYP2C19 gene polymorphism
  • Method and kit for detecting CYP2C19 gene polymorphism
  • Method and kit for detecting CYP2C19 gene polymorphism

Examples

Experimental program
Comparison scheme
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Embodiment 1

[0129] Example 1 Kit

[0130] This embodiment provides a detection kit for detecting CYP2C19 gene polymorphism, the composition of the detection kit of the present invention is as follows:

[0131]

Embodiment 2

[0132] Example 2 Detection method for detecting CYP2C19 gene polymorphism

[0133] Step 1: Preparation of PCR reaction system

[0134] Take out CYP2C19 wild-type PCR reaction solution A, CYP2C19 mutant PCR reaction solution A and CYP2C19 PCR reaction solution B from the kit, thaw at room temperature, shake and mix well, and centrifuge briefly at 8,000rpm before use.

[0135] The CYP2C19 reagent single serving wild-type PCR reaction tube system is prepared as follows:

[0136]

[0137] The preparation of CYP2C19 reagent single mutant PCR reaction tube system is as follows:

[0138]

[0139] The components were thoroughly mixed to prepare corresponding CYP2C19 wild-type PCR reaction tubes and CYP2C19 mutant PCR reaction tubes, centrifuged briefly, and 20 μl of the amplification system was dispensed into PCR tubes.

[0140] Step 2: Process the sample to be tested

[0141] Directly use 30 cases of whole blood samples to be tested as templates or use nucleic acid extractio...

Embodiment 3

[0148] Example 3 Detection of CYP2C19 Gene Polymorphism Using Human Clinical Whole Blood Samples as a Template

[0149] In this embodiment, human clinical whole blood samples are used as a template to detect CYP2C19*2, CYP2C19*3, and CYP2C19*17 gene polymorphisms according to the kit of the present invention.

[0150] This embodiment adopts the PCR reaction system and PCR reaction conditions that have been described in this specification, and the specific operation method is the same as that of Embodiment 2. In this embodiment, 15 representative human clinical whole blood samples were selected, and further, 15 cases of whole blood samples were directly added to PCR reaction wells for detection on the machine. By comparing the genotyping results with the gold standard sequencing, it was found that 14 cases of CYP2C19*1 / *1, 2 cases of CYP2C19*1 / *2, 2 cases of CYP2C19*1 / *3, and CYP2C19*1 / * were detected at the same time 2 cases of type 17, the representative genotyping results a...

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Abstract

The invention provides a method and a kit for detecting CYP2C19 gene polymorphism,specifically, the ARMS-PCR method is combined with an added PCR amplification blocker, and the kit for detecting the CYP2C19 gene polymorphism and with high specificity, high sensitivity, high flux and simple operation is provided. Through development and design of ARMS primers combined with the PCR amplification blocker, the specificity and accuracy are greatly improved. The invention provides a primer and probe for detecting CYP2C19 gene polymorphism and with high sensitivity and specificity, and provides the kit and a method for detecting the CYP2C19 gene polymorphism and with simple operation, accurate results and convenient popularization.

Description

technical field [0001] The invention belongs to the field of biotechnology and detection, in particular, the invention relates to a method and kit for detecting CYP2C19 gene polymorphism Background technique [0002] Cytochrome P450 (cytochrome P450, referred to as CYP450) is a superfamily of heme-thiolate proteins, which participate in the metabolism of endogenous substances and exogenous substances including pharmaceutical environmental compounds. CYP2C19 is one of the most important drug-metabolizing enzymes in the CYP450 family, and its genetic polymorphism affects the metabolism of many drugs such as clopidogrel, omeprazole, phenytoin and diazepam. The CYP2C19 gene is located on the q24.1-24.3 zone of human chromosome 10, and the wild type is CYP2C19*1 / *1. Gene mutations are common in the Chinese population, and the common mutation types are CYP2C19*2 (exon 5 G681A) and CYP2C19*3 (exon 4 G636A). In 2006, Sim et al. discovered a new CYP2C19 allele, CYP2C19*17, with the...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883C12Q1/686C12N15/11
CPCC12Q1/686C12Q1/6883C12Q2600/106C12Q2600/156C12Q2563/107C12Q2545/114
Inventor 蒋析文刘悦朱小亚黄志文
Owner DAAN GENE CO LTD
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