Primer set capable of synchronously detecting related gene polymorphism of anticoagulant drug and application

A gene polymorphism and drug technology, applied in the direction of recombinant DNA technology, microbial measurement/testing, biochemical equipment and methods, etc., can solve problems affecting drug efficacy and adverse reactions, reduce amplification costs, and ensure reliability Sexual, high-sensitivity effects

Inactive Publication Date: 2018-04-13
KUNMING UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0011] Polymorphisms leading to loss of CYP2D6 enzyme activity can affect antipyrine, codeine, beta-blockers such as metoprolol and carvedilol, clomipramine, nortriptyline, desipa In vivo metabolism of doxepin, doxepin, imipramine,...

Method used

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  • Primer set capable of synchronously detecting related gene polymorphism of anticoagulant drug and application
  • Primer set capable of synchronously detecting related gene polymorphism of anticoagulant drug and application
  • Primer set capable of synchronously detecting related gene polymorphism of anticoagulant drug and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Example 1: VKORC1-1639 G>A, CYP4F2*3, CYP2C9*3, CYP2C19*2, CYP2C19*3, CYP2D6* 10 Gene polymorphism SNaPshot detection

[0036] This detection method is to amplify the gene fragment where the SNP site to be detected is amplified by multiplex PCR, and then use fluorescent labeling single base extension technology (SNaPshot) combined with capillary electrophoresis technology to detect gene polymorphism, and the detection results are analyzed by GENEMAPPER software. The SNP site corresponding to the extension product is determined according to the shift position of the peak, and the genotype of the SNP site is determined according to the fluorescence signal of the peak.

[0037] This detection method can be used to detect DNA samples derived from EDTA anticoagulated peripheral blood. After the multiplex PCR method is used to amplify the gene fragment where the SNP site to be detected is located, the fluorescent-labeled single-base extension technology (SNaPshot) combined ...

Embodiment 2

[0051] (1) Primer specificity

[0052] Each primer was blasted with UCSC, the results are as follows: VKORC1-1639 G>A The amplified fragment is located at chr1631096202-31096491, with a length of 290bp; CYP4F2 *3 The amplified fragment is located at chr1915879539-15879818, with a length of 280bp; CYP2C9*3 The amplified fragment is located at chr1094981180-94981427, with a length of 248bp; CYP2C19*2 The amplified fragment is located at chr1094781745-94781964, with a length of 220bp; CYP2C19*3 The amplified fragment is located at chr1094780531-94780850, with a length of 320bp; CYP2D6*10 The amplified fragment is located at chr2242130632-42130878 with a length of 247bp.

[0053] The amplification results are shown in Table 4, and the amplified fragments of all primers covered the corresponding detection sites VKORC1-1639 G>A, CYP4F2*3, CYP2C9*3, CYP2C19*2, CYP2C19*3, CYP2D6*10 , no other homologous fragments.

[0054] Table 4 Primer Amplified Fragment

[0055] ...

Embodiment 3

[0057] Example 3: Specificity and Sensitivity Detection

[0058] 1. Specificity of primer combination detection

[0059] The specificity of this detection method is defined as the negative coincidence rate. The SnaPshot method was used to detect 30 samples of patients who needed to use anticoagulant drugs in the Department of Cardiology. The test results showed that: VKORC1 -1639 G>A negative rate is 80%, CYP4F2 *3 The negative rate was 83.3%, CYP2C9*3 The negative rate was 80%, CYP2C19 *2 The negative rate is 40%, CYP2C19*3 The negative rate was 90%, CYP2D6* 10 The negative rate was 33.3%. Among them, only 2 cases were negative for all sites, which was consistent with the test results of Sanger sequencing method (as shown in Table 5), and the detection specificity of the primer combination of the present invention was 100%.

[0060] Table 5 Primer Combination Detection Specificity

[0061] .

[0062] 2. The sensitivity of the detection method

[0063] The sens...

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Abstract

The invention discloses a primer set capable of synchronously detecting related gene polymorphism of an anticoagulant drug. The primer set comprises a PCR amplification primer and a SNaPshot PCR primer; the detected sites include VKORC1-1639>A, CYP4F2*3, CYP2C9*3, CYP2C19*2, CYP2C19*3 and CYP2D6*10. Compared with a general method for individually amplifying a plurality of sites, the primer used for detecting has the advantages that the amplification processes are decreased, and the amplification cost is reduced; ,moreover, the high sensitivity, the high accuracy and the high precision can be ensured, the method reliability is ensured, and a doctor can accurately select drugs for patients and reasonably adjust the dosage of the drug.

Description

technical field [0001] The invention belongs to the field of gene polymorphism detection, in particular to a simultaneous detection VKORC1-1639 G>A, CYP4F2*3, CYP2C9*3, CYP2C19*2, CYP2C19*3, CYP2D6*10 Gene polymorphism primers and applications. Background technique [0002] One of the reasons for the high incidence of cardiovascular diseases is the formation of thrombus. Anticoagulant drugs and antiplatelet drugs are two types of drugs widely used in antithrombotic drugs, which play a leading role in the treatment of cardiovascular diseases. The treatment of cardiovascular and cerebrovascular thrombosis and the These two types of antithrombotic drugs are required in the interventional treatment of vascular diseases, and the two most commonly used drugs in clinical practice are warfarin and clopidogrel. Based on its antithrombotic mechanism of action, if used improperly, it may lead to bleeding risks and endanger the lives of patients. Both warfarin and clopidogrel nee...

Claims

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Application Information

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IPC IPC(8): C12Q1/6883C12Q1/6858C12N15/11
CPCC12Q1/6858C12Q1/6883C12Q2600/106C12Q2600/136C12Q2600/156C12Q2600/16C12Q2537/143C12Q2531/113
Inventor 唐文如罗瑛旦菊花刘宁盛苗苗李静怡念孙琪王思皓张继虹贾舒婷吴晓明刘静谢晓丽周若宇
Owner KUNMING UNIV OF SCI & TECH
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