ARMS primer for detecting CYP2C9 gene polymorphism

A gene polymorphism and gene technology, applied in the field of life science and biology, can solve the problems of ineffective amplification, high false positive rate of hybridization chip detection method, high annealing temperature, expensive instruments and equipment, etc., to improve detection accuracy and reliability performance, expanding the optimum annealing temperature range, and avoiding false positives

Inactive Publication Date: 2017-02-15
SHANGHAI PERSONAL BIOTECH
View PDF1 Cites 2 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Among them, DNA sequencing is the gold standard for genotyping, but this method has some limitations in clinical application: low detection sensitivity, time-consuming, complicated operation, easy contamination, low throughput, etc.
Restriction fragment length polymorphism is only applicable when there is an appropriate specific restriction endonuclease recognition sequence near the mutation site, and its application is limited
The h

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • ARMS primer for detecting CYP2C9 gene polymorphism
  • ARMS primer for detecting CYP2C9 gene polymorphism
  • ARMS primer for detecting CYP2C9 gene polymorphism

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0041] Embodiment 1 A kind of CYPC2C9 genotyping detection based on ARMs-PCR method

[0042] PCR Amplification Reaction Solution The amplification reaction solution is 20ul, and the Q5hifi DNA pilymerase reagent kit from NEB Company is used, including 5xQ5buffer, 5xGC buffer, dNTP, Q5hifi DNA pilymerase and 5xGC enhancer. There are 2 PCR reaction tubes per person, and tubes 1 and 2 are specific reaction tubes. See Table 1 for details

[0043] Table 1

[0044]

Embodiment 2

[0046] A kind of operation steps of CYPC2C9 genotyping detection based on ARMs-PCR method

[0047] (1) Extract the whole blood DNA or saliva DNA of the subject, and dilute it into a 20ng / ul DNA solution;

[0048] (2) PCR amplification: detection is carried out on a conventional PCR instrument, and the reaction conditions are shown in Table 2:

[0049] Table 2

[0050]

[0051] See Table 3 for the preparation method of PCR amplification system reagents:

[0052] table 3

[0053] 5x Q5buffer 4ul 5x GC buffer 4ul 10mM dNTPs 0.4ul Q5hifi DNA pilymerase 0.2ul 5xGC enhancer 4ul template DNA 20ng 10uM Forward Primer 1ul 10uM Reverse Primer 1ul Nuclease-free water up to 20ul

[0054] (3) Electrophoresis: 2% agarose gel electrophoresis, 110V, 25min. Take pictures in the gel imaging system to obtain the electrophoresis results. According to the presence or absence of the target band, perform genotype analysis.

[...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention discloses an ARMS primer for detecting CYP2C9 gene polymorphism. Aiming at sequence changes in every SNP of CYPC2C9*2(rs1799853, C430T, Arg144Cys) and CYP2C9*3(rs1057910, A1075C, Ile359Leu), the specificity ARMS primer and shared primers are designed. Mismatched base group (base group is marked with underline) is introduced to the second or third place of the 3' terminal end of the specificity ARMS primer so as to enlarge efficiency gap generating when specificity ARMS primer 3' terminal base are complementary and are not complementary to the SNP locus base on the DNA template, thus sharply improving specificity and accuracy of the detection.

Description

technical field [0001] The invention belongs to the field of life science and biotechnology, and in particular relates to ARMS primers for detecting mutations of exons 3 and 7 of CYP2C9 gene, which can be used for rapid detection of mutations of polymorphic sites of CYP2C9 gene by using ordinary PCR technology and gel electrophoresis. Background technique [0002] Human gene polymorphisms play an important role in elucidating the susceptibility and tolerance of the human body to diseases and poisons, the diversity of clinical manifestations of diseases, and the responsiveness to drug treatment. [0003] The CYP2C9 gene is located on human chromosome 10, with a total length of 83529bp and a total of 10 exons. CYP2C9 is an important member of the second subfamily of cytochrome P450 enzymes (CYP), accounting for 20% of the total amount of P450 proteins in liver microsomal. CYP2C9 participates in the hydroxylation metabolism of various drugs such as anticoagulant drugs, anticon...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6858C12Q2531/113C12Q2535/137
Inventor 丁慧朱月艳孙子奎
Owner SHANGHAI PERSONAL BIOTECH
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap