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Detection kit for metabolic markers of ondansetron and tropisetron, detection method of detection kit and application of detection kit and detection method

A technology for detection kits and metabolic markers, which is used in the determination/inspection of microorganisms, biochemical equipment and methods, DNA/RNA fragments, etc., which can solve the problems of long detection cycle, easy contamination of amplified products, and complicated operation steps. question

Pending Publication Date: 2021-12-07
上海普然生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Among them, the sequencing method and the chip method have cumbersome operation steps, long detection cycle, and the amplification product is prone to contamination; the high-resolution melting curve method has simple steps, low specificity, and high requirements for equipment; The specific amplification method uses ARMS primers for specific amplification, and the design of the primers is difficult to optimize, and the detection conditions are strict.
Taqman fluorescent probe method has high test cost, and the amplification throughput of multiple genes is not high

Method used

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  • Detection kit for metabolic markers of ondansetron and tropisetron, detection method of detection kit and application of detection kit and detection method
  • Detection kit for metabolic markers of ondansetron and tropisetron, detection method of detection kit and application of detection kit and detection method
  • Detection kit for metabolic markers of ondansetron and tropisetron, detection method of detection kit and application of detection kit and detection method

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Experimental program
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Effect test

Embodiment 1

[0046] Embodiment 1, the preparation of kit

[0047] The rapid reaction kit of the present invention designs specific amplification primers and sequencing primers for CYP2D6 (C100T), CYP2D6 (G1846A) and CYP2D6*5 for amplification and pyrosequencing detection. Designing primers based on rapid amplification technology is one of the keys of the present invention. Gene polymorphism sequence is subject to the public sequence in Genebank.

[0048] (1) The primer sequences of this embodiment are as follows:

[0049]

[0050]

[0051] The copy number detection fragment of CYP2D6*5 is located in exon 9 of the CYP2D6 gene. Through the same sequence as CYP2D6 and CYP2D8 but different from CYP2D7, the related fragments of CYP2D6 and CYP2D8 genes are amplified at the same time, and then the PCR amplification product is used for Pyrosequencing. Since there are only two copies of CYP2D8, the peak height ratios of different bases at the same position in the measured sequence of the C...

Embodiment 2

[0059] Embodiment 2, pyrophosphate detection

[0060] The instruments adopted in the present invention are as follows: amplification instrument, pyrosequencer (Wuhan First Biotechnology Co., Ltd.).

[0061] (1) Reagent preparation (reagent preparation room)

[0062] Take out the reagents in advance, vortex the PCR reaction solution for 15 seconds, and centrifuge at low speed for later use. Determine the number of reactions N, N = number of samples to be tested (n) + number of quality control products (1) + blank control. It is recommended to conduct positive control and blank control analysis for each PCR experiment at the same time. Then the reaction solution was dispensed into PCR reaction tubes at 16 μL / tube.

[0063] (2) Sample testing (sample preparation room)

[0064] Add EDTA anticoagulated whole blood, positive control and blank control into the PCR reaction tube according to the sample volume of 4 μL, close the tube cap tightly, centrifuge at low speed for 15 seco...

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Abstract

The invention discloses a detection kit for metabolic markers of ondansetron and tropisetron, a detection method of the detection kit and application of the detection kit and the detection method. The detection kit is used for detecting gene polymorphism of the metabolic markers CYP2D6C100T, CYP2D6G1846A and CYP2D6*5 of the ondansetron and the tropisetron. According to the kit, specific amplification primers and sequencing primers are designed according to the polymorphism of the three genes CYP2D6C100T, CYP2D6G1846A and CYP2D6*5. The kit comprises the following components of an amplification reaction solution, the CYP2D6C100T sequencing primer, the CYP2D6G1846A sequencing primer, the CYP2D6*5 sequencing primer and a positive control. According to the invention, blood direct expansion, rapid amplification and optimized pyrosequencing technologies are combined to detect gene polymorphism related to prediction of drug effects and adverse reactions of the ondansetron and the tropisetron, and suggestions from the gene perspective are provided for clinical use of the ondansetron and the tropisetron.

Description

technical field [0001] The invention relates to a detection kit for ondansetron and tropisetron metabolic markers, a detection method and application thereof, and belongs to the field of gene detection. Background technique [0002] Postoperative nausea and vomiting (PONY) occurs in 20% of hospitalized surgical patients, and 80% of high-risk PONV patients. At the same time, PONV can also induce related complications, including dehydration, electrolyte disturbance, incision dehiscence, bleeding, and aspiration pneumonia. Ondansetron and tropisetron are highly selective 5-hydroxytryptamine receptor antagonists, which are used to prevent and treat postoperative nausea and vomiting caused by chemotherapy and radiotherapy. However, their curative effect is not satisfactory in some patients. [0003] CYP2D6 is one of the main drug-metabolizing enzymes of ondansetron and one of the important members of the CYP enzyme family. Although it only accounts for 2% and 9% of the total liv...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883C12Q1/686C12Q1/6869C12N15/11
CPCC12Q1/6883C12Q1/686C12Q1/6869C12Q2600/106C12Q2600/156C12Q2537/143C12Q2565/301
Inventor 周虹桥刘丹孙悦
Owner 上海普然生物科技有限公司
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