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Primer composition for distinguishing individualized medication type of valsartan

A technique of primer composition and primer combination, which is applied in the field of molecular biology detection and can solve problems such as limitations

Inactive Publication Date: 2020-05-05
BIOYONG TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the method and kit provided by this invention patent can only detect polymorphic sites on the homocysteine ​​metabolism pathway, and cannot detect polymorphic sites not involved in homocysteine ​​metabolism , has limitations in clinical medication guidance

Method used

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  • Primer composition for distinguishing individualized medication type of valsartan
  • Primer composition for distinguishing individualized medication type of valsartan
  • Primer composition for distinguishing individualized medication type of valsartan

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0084] Example 1: Primer Design and Synthesis

[0085] The target SNP surrounding sequences of this kit are queried in db_SNP (build 132) and Hapmap (Rel 28, Phase II+III, Aug 10) databases, and use these sequences to design multiple PCR primers and single-base extension primers.

[0086] Corresponding specific PCR primer core sequences (SEQ1a to SEQ3a) and specific extension primer core sequences (SEQ1b to SEQ3b) were designed for the polymorphic sites of rs1799983, rs1042713 and rs1801253 related to discriminating drug types. The 3 pairs of PCR primers and the 3 extension primers constituted 3 independent reaction systems: SEQ1a / b constituted the first reaction system, and SEQ2a / b to SEQ3a / b constituted the second reaction system. In these two independent reaction systems, SEQ1a, SEQ2a to SEQ3a participate in two independent multiplex PCR reactions, respectively, and SEQ1b, SEQ2b to SEQ3b participate in two subsequent independent single-base extension reactions respectively....

Embodiment 2

[0088] Example 2: Sample DNA extraction

[0089] A total of 10 DNA samples from ordinary Chinese were collected and marked as A1-A10. Among them, human venous blood was collected according to the instructions for sample collection, DNA extraction, etc., and collected with an EDTA anticoagulant tube. According to the requirements of the instructions, the collected blood should not be stored at 2-8°C for more than a week, and at -20°C should not be stored for more than one month. It can be transported in a curling ice box or a foam box with ice sealed. It is recommended to use fresh blood as much as possible. Genomic DNA extraction. Since this kit does not provide human genomic DNA extraction reagents, commercial nucleic acid extraction kits (such as DNeasy Blood and Tissuekit from QIAGEN Company) were used to extract human genomic DNA from 200 μL of whole blood of each patient, with NanoDrop 2000 ( Thermo Co.) quantified and standardized to 30 ng / μL (A1-A10, respectively). A...

Embodiment 3

[0090] Example 3: Biological Experiment

[0091] Using ABI 9700 type PCR machine, according to the instructions, the two polymorphic loci related to the discriminating drug type were tested.

[0092] The components in the kit for PCR, PCR product purification and single base extension are:

[0093] serial number component name main ingredient Specification 1 PCR primer mix PCR primers 24μl / tube x1 tube 2 PCR reaction solution Taq enzyme, dNTP 72μl / tube x1 tube 3 Enzyme cleavage reaction solution SAP enzyme 48μl / tube x1 tube 4 Extension primer mix extension primer 24μl / tube x1 tube 5 extension reaction solution Single base elongase, ddNTP 24μl / tube x1 tube 6 Positive control Human genomic DNA (30ng / μL) 10μL / tube x1 tube

[0094] The concentration of each primer pair was 500 nmol / L.

[0095] According to the manual, the specific operation method is as follows:

[0096] 1. PCR amplification

[0097...

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Abstract

The invention provides a primer composition for distinguishing an individualized medication type of an antihypertensive drug valsartan. A method for distinguishing individualized medication type of valsartan comprises the following steps: respectively designing multiple amplification primers and extension primers according to multiple target SNP sites to be detected; preparing a multiple amplification primer reaction system and an extension primer reaction system; in a reaction system, using multiple sets of primers to simultaneously perform amplification and single base extension reactions onthe multiple target SNP sites respectively; performing time-of-flight mass spectrometry analysis on the products after the single base extension reaction, identifying genotypes of different drug metabolism related SNPs according to the products of different molecular weight extension primers represented by mass spectrum peaks, and guiding the individualized medication of the antihypertensive drugvalsartan. The primer composition can be used for simultaneously detecting the SNP sites related to metabolism of the antihypertensive drug valsartan, and has advantages of low cost, no need of probesynthesis, short time consumption, simple and convenient result analysis and extremely wide application field.

Description

technical field [0001] The invention belongs to the field of molecular biology detection, and relates to a method for detecting multiple PCR single-base extension products using mass spectrometry characteristic peaks and the product thereof. The method can simultaneously detect multiple PCR single-base extensions in two multiplex PCR reactions Amplified oligonucleotide product. More specifically, the method utilizes different time-of-flight mass spectrometry characteristic peaks generated by different target oligonucleotide fragments in the process of mass spectrometry typing, and simultaneously detects multiple target SNP sites to guide the antihypertensive drug valsa. individualized medicine of tannin. Background technique [0002] Human genetic information is stored in the genome. In 2002, the final completion of the Human Genome Project, an international cooperation project, drew a fine map of the structure of the human genome and provided a reference sequence for relat...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883C12N15/11C12Q1/6858
CPCC12Q1/6858C12Q1/6883C12Q2600/106C12Q2600/156C12Q2600/16C12Q2531/113C12Q2537/143C12Q2533/101C12Q2565/627
Inventor 马庆伟张海燕钟逾刘昕超
Owner BIOYONG TECH
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