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Primer, molecular beacon, kit and detection method for CYP2C19*3 gene polymorphism rapid detection

A CYP2C19, 1.CYP2C19 technology, applied in the field of primers for the rapid detection of CYP2C19*3 gene polymorphisms, can solve the problems of increasing the risk of disease treatment for patients and difficult to meet the rapid diagnosis of clinical diseases

Inactive Publication Date: 2017-11-03
重庆京因生物科技有限责任公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, it is difficult to meet the rapid diagnosis of clinical diseases, which increases the risk of patients' disease treatment

Method used

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  • Primer, molecular beacon, kit and detection method for CYP2C19*3 gene polymorphism rapid detection
  • Primer, molecular beacon, kit and detection method for CYP2C19*3 gene polymorphism rapid detection
  • Primer, molecular beacon, kit and detection method for CYP2C19*3 gene polymorphism rapid detection

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 3

[0079] (3) Preparation of 200X cell lysate in embodiment three

[0080] Using a 1.5 mL centrifuge tube, add 300 μL sodium dodecyl sulfate and 56.34 μL polyethylene glycol octylphenyl ether, and then add 643.66 μL nuclease-free water to a total volume of 1000 μL to make the sodium dodecyl sulfate When the final concentration reaches 3%, the final concentration of polyethylene glycol octylphenyl ether reaches 6%, which is 200X cell lysate, then shakes and mixes well, and stores at -4°C.

[0081] The gene sequence of the forward primer 5'-3' is: CAGCAATTTCTTAACTTGATGGA;

[0082] The gene sequence of the reverse primer 5'-3' is: CAATATAGAATTTTGGATTTCCCAG.

[0083] Such as figure 1 Shown: figure 1 In, Molecular Beacon: molecular beacon; Target: target sequence; Hybrid: hybridization; Fluorophore: fluorescent molecule; Quancher: quencher molecule

[0084] The gene sequence of wild-type probe 5'-3' is: (Fam mark)-CGAGCCTAAGCACCCCCTGGATCCGGC

[0085] TCG-(BHQ1 marker);

[0086] ...

Embodiment 4

[0115] Compared with Example 2, Example 4 is only different in the amount of PCR reaction solution, specific primers and molecular beacons used, and the genotype of CYP2C19*3 can also be obtained from the test results. It can be seen that the CYP2C19 of the present invention *3 The success of rapid detection of polymorphisms is not closely related to the amount of PCR reaction solution, specific primers and molecular beacons used.

[0116] Depend on Figure 14 , Figure 15 and Figure 16 It can be seen that only the "G" line has exponential growth, while the "A" line has no exponential growth, so the test result is wild homozygous "GG";

[0117] Depend on Figure 17 , Figure 18 and Figure 19 It can be seen that both the "G" line and the "A" line have exponential growth, indicating that the test result is a mutation heterozygous "GA";

[0118] Depend on Figure 20 , Figure 21 and Figure 22 It can be seen that only the "A" line has exponential growth, while the "G" ...

Embodiment 5

[0119] The difference between Example 5 and Example 2 lies in the number of swabs. It can be concluded that the genotype of CYP2C19*3 can be obtained when the number of swabs is 3, 5 or 7.

[0120] The nucleotide sequence list is as follows:

[0121] Chongqing Jingyin Biotechnology Co., Ltd.

[0122] Primers, molecular beacons, kits and detection methods for rapid detection of CYP2C19*3 gene polymorphism

[0123] 4

[0124] 1

[0125] 23

[0126] DNA

[0127] Artificial sequence

[0128]

[0129] prim_bind

[0130] 1

[0131] CAGCAATTTCTTAACTTGATGGA

[0132] 2

[0133] 25

[0134] DNA

[0135] Artificial sequence

[0136]

[0137] prim_bind

[0138] 2

[0139] CAATATAGAATTTTGGATTTCCCAG

[0140] 3

[0141] 30

[0142] RNA

[0143] Artificial sequence

[0144]

[0145] misc_binding

[0146] 3

[0147] CGAGCCTAAGCACCCCCTGGATCCGGCTCG

[0148] 4

[0149] 30

[0150] DNA

[0151] Artificial sequence

[0152]

[0153] misc_binding

[0154] 4 ...

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Abstract

The invention discloses a kit for CYP2C19*3 gene polymorphism rapid detection. The kit is characterized by comprising a PCR reaction liquid, a specificity primer and a molecular beacon; the PCR reaction liquid comprises the following raw materials and concentrations: DNA polymerase 0.05 to 0.12 U / [mu]L; dNTPs 0.2 mM; 5X reaction buffer 1X; MgCl2 1.5 to 3.5 mM; lauryl sodium sulfate 0.0005 to 0.015 %(w / v); polyethylene glycol octylphenol ether 0.001 to 0.03 %(w / v); the specificity primer comprises a positive primer and a negative primer, and the final concentration of the positive primer and the final concentration of the negative primer are 0.2 to 0.5 [mu]M respectively; the molecular beacon comprises a mutant type probe and a wild type probe, and the final concentration of the mutant type probe and the final concentration of the wild type probe are 0.4 to 0.6 [mu]M; and the kit is used for detecting cell samples. The technical problem to be solved by the invention is to provide a primer, a molecular beacon, the kit and a detection method for CYP2C19*3 gene polymorphism rapid detection, which are simple in operation and rapid in detection and avoid pollution.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to a primer, a molecular beacon, a kit and a detection method for rapid detection of CYP2C19*3 gene polymorphism. Background technique [0002] Nucleotide polymorphism (Single Nucleotide Polymorphism, SNP) mainly refers to the DNA sequence polymorphism caused by the variation of a single nucleotide at the genome level. SNPs exist widely in the human genome, with an average of 1 SNP per 1000 base pairs. These genetic variations underlie human diversity, and some of them are intimately associated with susceptibility to many genetic diseases and interindividual differences in drug response. Therefore, SNP has important application value in analysis and diagnosis, clinical testing, forensic science, pathogen detection, genetic disease and new drug development, etc.; [0003] CYP2C19 is an important member of the cytochrome 450 enzyme (CYP450) family, involved in the metabolism of a va...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6876C12Q2600/106C12Q2600/156
Inventor 吴丹贺庭祯罗德朋黄东旗向·霄熊伟
Owner 重庆京因生物科技有限责任公司
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