Primer composition and kit for detecting sulfonylurea-related gene

A technology of primer composition and sulfonylureas, which is applied in the direction of recombinant DNA technology, microbial measurement/testing, biochemical equipment and methods, etc., can solve the problems of high price and limitation, and achieve simple method, rapid detection and high sensitivity high effect

Inactive Publication Date: 2018-09-14
宁波美丽人生医学检验所有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, similar kits currently on the ma...

Method used

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  • Primer composition and kit for detecting sulfonylurea-related gene
  • Primer composition and kit for detecting sulfonylurea-related gene
  • Primer composition and kit for detecting sulfonylurea-related gene

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0071] 1. Sample processing and nucleic acid extraction:

[0072] Use a commercial DNA extraction kit to process the sample. For specific operations, refer to the kit instruction manual. The concentration and purity of the extracted DNA need to be measured with a UV spectrophotometer. The DNA OD 260 / OD 280 The value of DNA should be between 1.8 and 2.0, and the concentration should be between 5 and 50 ng / μL. If the DNA quality of the sample is unqualified, it should not be used for detection. If it is lower than 5 ng / μL, the nucleic acid should be extracted again, and if it is higher than 50 ng / μL, it should be properly diluted. To the specified concentration range, the extracted DNA should be detected immediately, otherwise it should be stored below -20°C, and the storage time should not exceed 6 months. The quality control product of the kit was thawed at room temperature before use, vortexed for 10 seconds, and centrifuged at 2000rpm for 15 seconds before use.

[0073] 2...

Embodiment 2

[0092] The primer composition for detecting sulfonylurea drug-related genes includes a CYP2C9 primer set, a KCNJ11 primer set and an ABCC8 primer set;

[0093] The CYP2C9 primer set includes the following primers:

[0094] P1-FW: 5'-AGAGGAGCATTGAGGAAC-3';

[0095] P1-FS: 5'-AAGAGGAGCATTGAGGACT-3';

[0096] P1-R: 5'-TAACAACCAGGACTCAT-3';

[0097] P2-FW: 5'-GCACGAGGTCCAGAGATAGA-3';

[0098] P2-FS: 5'-GCACGAGGTCCAGAGATAGC-3';

[0099] P2-R: 5'-CGGTGATGGTAGAGGTTTAA-3';

[0100] The KCNJ11 primer set includes the following primers:

[0101] P3-F: 5'-GCACGGTACCTGGGCAC-3';

[0102] P3-RW: 5'-GCACGGTACCTGGGCAT-3';

[0103] P3-RS: 5'-CACCGAGAGGACTCTGCA-3';

[0104] The ABCC8 primer set includes the following primers:

[0105] P4-FW: 5'-CGTCAATGCCCTCATGT-3';

[0106] P4-FS: 5'-CGTCAATGCCCTCATGT-3';

[0107] P4-R: 5'-ACTGCGATGTCTGAATA-3'.

[0108]The above primer composition is used in the preparation of reagents for detecting gene polymorphisms related to sulfonylurea drugs....

Embodiment 3

[0111] The primer composition for detecting sulfonylurea drug-related genes includes a CYP2C9 primer set, a KCNJ11 primer set and an ABCC8 primer set;

[0112] The CYP2C9 primer set includes the following primers:

[0113] P1-FW: 5'-AGAGGAGCATTGAGGAAC-3';

[0114] P1-FS: 5'-AAGAGGAGCATTGAGGACT-3';

[0115] P1-R: 5'-TAACAACCAGGACTCAT-3';

[0116] P2-FW: 5'-GCACGAGGTCCAGAGATAGA-3';

[0117] P2-FS: 5'-GCACGAGGTCCAGAGATAGC-3';

[0118] P2-R: 5'-CGGTGATGGTAGAGGTTTAA-3';

[0119] The KCNJ11 primer set includes the following primers:

[0120] P3-F: 5'-GCACGGTACCTGGGCAC-3';

[0121] P3-RW: 5'-GCACGGTACCTGGGCAT-3';

[0122] P3-RS: 5'-CACCGAGAGGACTCTGCA-3';

[0123] The ABCC8 primer set includes the following primers:

[0124] P4-FW: 5'-CGTCAATGCCCTCATGT-3';

[0125] P4-FS: 5'-CGTCAATGCCCTCATGT-3';

[0126] P4-R: 5'-ACTGCGATGTCTGAATA-3'.

[0127] The primer composition also includes an internal control primer set, which includes the following primers:

[0128] CF: 5'-AGCAAG...

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Abstract

The invention relates to a primer composition and a kit for detecting sulfonylurea-related genes, wherein the primer composition comprises a CYP2C9 (cytochrosome P4502C9) primer set, a KCNJ11 primer set and a ABCC8 primer set, and the kit comprises the above-mentioned primer compositions. Compared with the prior art, with ARMS (the amplification refractory mutation system) technology combined withSYBR dye, the kit for rapidly, sensitively and simply detecting the polymorphisms of sulfonylurea-related genes such as CYP2C9, KCNJ11 and ABCC8 is developed, and the kit comprises a specific ARMS detection primer, an internal control primer and aPCR reaction solution. By designing ARMS primers and replacing Scorpions probes with SYBR dyes, the primer composition and the kit for detecting the sulfonylurea-related genes have the advantages of greatly reducing detection costs, being suitable for for detection of CYP2C9, KCNJ11 and ABCC8 gene polymorphisms in Chinese patients, being rapid in detection, high in sensitivity, strong in specificity, simple in method and accurate in results, and being suitable for popularization and application.

Description

technical field [0001] The invention belongs to the technical field of gene detection, and relates to a primer composition and a kit for detecting genes related to sulfonylurea drugs. Background technique [0002] Sulfonylureas (SU), one of the main oral drugs for the treatment of type 2 diabetes, close the potassium channels of pancreatic β cells by binding to sulfonylurea receptors, thereby stimulating insulin secretion. Studies have found that there are large individual differences in the efficacy of sulfonylureas. About 10%-20% of patients have higher efficacy than the average level, but 10%-20% of patients still fail to treat after initially taking sulfonylureas. Studies have pointed out that this phenomenon is mostly due to the mutation of Kir6.2 and SUR1 receptors that act on the KATP channel, encoded by KCNJ11 and ABCC8, respectively, and CYP2C9 that metabolizes sulfonylureas. [0003] CYP2C9 is a common drug enzyme metabolism gene in the CYP450 family, and the most...

Claims

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Application Information

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IPC IPC(8): C12Q1/6883C12Q1/6858C12N15/11
CPCC12Q1/6858C12Q1/6883C12Q2600/106C12Q2600/156C12Q2563/107C12Q2535/137C12Q2531/113
Inventor 田晓丽
Owner 宁波美丽人生医学检验所有限公司
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