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Primer probe combination, kit and method for detecting copy number variation and genotyping of human CYP2D6

A copy number variation and genotyping technology, applied in biochemical equipment and methods, recombinant DNA technology, and microbial assay/inspection, etc. The experimental steps are cumbersome and other problems, to achieve the effect of preventing false positive amplification, excellent detection limit, and reducing toxic and side reactions

Pending Publication Date: 2021-07-30
上海康黎诊断技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] The purpose of the present invention is to provide a primer probe combination, kit and method for human CYP2D6 copy number variation and genotyping detection, thereby solving the detection reagent cost in the prior art for human CYP2D6 copy number variation and genotyping detection High, low accuracy, high requirements for determination of instrument performance and result interpretation threshold, cumbersome experimental steps

Method used

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  • Primer probe combination, kit and method for detecting copy number variation and genotyping of human CYP2D6
  • Primer probe combination, kit and method for detecting copy number variation and genotyping of human CYP2D6
  • Primer probe combination, kit and method for detecting copy number variation and genotyping of human CYP2D6

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Embodiment 1

[0034] Embodiment 1 Designs primers and probes

[0035] According to the present invention, the TaqMAN fluorescence quantitative method is used to target the CYP2D6 exon 9 region, excluding the identification of the copy number of *36 and *5 non-functional alleles, specific amplification, and adding double internal references TERT and RPPH as qualified evaluation samples parameters, to avoid the risk of misjudgment due to sample quality problems, and realize simultaneous detection of three target genes (2D6+TERT+RPPH) in a single tube. The present invention designs primers and probes for human CYP2D6 gene, TERT internal reference gene, and RPPH internal reference gene respectively. The nucleotide sequences thereof are shown in Table 1 below, and the preferred concentration of each primer and probe is also shown.

[0036] Table 1 The primer probe sequences designed for the detection of human CYP2D6 copy number

[0037]

[0038]

[0039] At the same time, the inventor als...

Embodiment 2

[0042] Example 2 A kit for human CYP2D6 copy number variation and genotyping detection

[0043] 1. Main components

[0044] According to this preferred embodiment, a kit for detecting copy number variation and genotyping of human CYP2D6 is provided, and the main components of the kit are shown in Table 3 below.

[0045] Table 3 The main components of the kit

[0046]

[0047] 2. Components that must be detected but not included in the kit

[0048] 1.5ml centrifuge tube (for configuring PCR reaction solution and DNA extraction), 0.2ml PCR tube or 8-tube strip or 96-well plate, tip with filter plug (1ml, 200μL and 10μL), DNA extraction kit (recommended Qiamp DNA Blood Mini Kit, Enrich Swab / Blood Spot Extraction Kit).

[0049] 3. Applicable instruments

[0050] Applicable to the Real-time PCR amplification instrument of ABI7500 model.

[0051] 4. Sample requirements

[0052] 4.1 Applicable samples are EDTA anticoagulated whole blood, oral swabs and blood cards; samples s...

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Abstract

The invention provides a primer probe combination, a kit and a method for detecting copy number variation and genotyping of human CYP2D6. The nucleotide sequences of the primer probe combination for detecting the copy number variation of the human CYP2D6 are as shown in SEQ ID No.1 to SEQ ID No.9, and the nucleotide sequences of the primer probe combination for detecting the genotyping of the human CYP2D6 are as shown in SEQ ID No.10 to SEQ ID No.17. The primer probe combination can be used for in-vitro qualitative detection of CYP2D6 copy number variation and * 10 and * 41 genotypes in human genome DNA, realizes simultaneous detection of * 10 and * 41 genotypes in a single tube on imported / domestic fluorescent quantitative PCR equipment without mutual interference, has the advantages of high accuracy, good stability and excellent detection limit, and has a wide application prospect. According to the detection result, clinical selection of antipsychotic drugs can be assisted to realize medication guidance for mental / depression patients, the curative effect is ensured, and toxic and side effects are reduced.

Description

technical field [0001] The invention relates to the field of molecular biology, and more specifically relates to a primer-probe combination, kit and method for human CYP2D6 copy number variation and genotyping detection. Background technique [0002] Fluorescent quantitative PCR technology is a simple, fast, highly sensitive, and specific method for gene detection. CYP2D6 enzymes are involved in the liver metabolism of about 25% of commonly used drugs, including antidepressants, antipsychotics, antiarrhythmics, opioids and Beta-blockers. Differences in CYP2D6 alleles will increase, decrease or inactivate the enzyme activity encoded by it, which may lead to differences in the risk of drug reactions and adverse reactions among individuals, while the copy number variation of the gene (such as *5 deletion, *36X2 , *36+*10 rearrangement, etc.) play a key role in their individual drug metabolism. Among them, *1 (34.9%), *2 (13.3%), *5 (5%), *10 (43%), *36 (1.2%), *41 (2.34%) are...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6858C12Q1/6883C12N15/11
CPCC12Q1/6858C12Q1/6883C12Q2600/106C12Q2600/156C12Q2600/166C12Q2545/101C12Q2545/113C12Q2547/101C12Q2531/113C12Q2563/107C12Q2537/16C12Q2521/531
Inventor 查广彬何炯张辉韩燕
Owner 上海康黎诊断技术有限公司
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