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Gene mutation detection primer and application

A base and base pairing technology, which is applied in the determination/inspection of microorganisms, biochemical equipment and methods, DNA/RNA fragments, etc., can solve the problems of weak detection ability, weak sensitivity, weak specificity, etc., and achieve inhibition The effect of nonspecific amplification

Active Publication Date: 2014-04-09
常州可尔生命科技有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The technical problem to be solved by the present invention is: aiming at the problems of weak sensitivity, weak detection ability, and weak specificity in the existing technology for detecting target gene mutations, to provide a gene mutation that can detect low-copy copies, and to provide a new Primers for detection of gene mutations in the detection system and their application, so that the target gene mutations that need to be detected can be amplified efficiently and specifically

Method used

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  • Gene mutation detection primer and application
  • Gene mutation detection primer and application
  • Gene mutation detection primer and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] For hepatitis B virus C gene detection, this example uses the method of the present invention to design and detect 180 drug resistance sites.

[0043] Material:

[0044] material name

company

Hotstart Taq polymerase

Shanghai Sangong

Mg2+

Shanghai Sangong

PCR buffer

Shanghai Sangong

dNTP mix (dA, dG, dC, dT)

Promega

[0045] 1. Sample and standard preparation:

[0046] Accept clinical samples, 200ul serum, add proteinase K and buffer AL, mix for 15s, digest at 56°C for 10 minutes, mix again, add 200ul absolute ethanol, mix for 15s, carefully transfer the liquid into a 2ml spin column, Centrifuge at 6000g (8000rpm) for one minute. Add 500ul buffer AW1, centrifuge at 6000g (8000rpm) for 1 minute, open the lid carefully and add 500ul AW2, centrifuge at 6000g (8000rpm) for 1 minute, centrifuge the empty tube at 20000g for 3 minutes, add 50ul ATE buffer and the center of the membrane, room temperature for 10 m...

Embodiment 2

[0068] Detection of lamivudine HBV rtL180M resistance site.

[0069] my country is the hardest-hit area of ​​hepatitis B virus infection. The current clinical treatment of chronic hepatitis B is to inhibit virus replication, normalize transaminases and improve liver tissue. The long-term effective method is the treatment of antiviral drugs. These drugs, such as lamivudine and adefovir, can effectively inhibit the replication of hepatitis B virus. However, due to the lack of a rigorous correction system for the transcription and replication of hepatitis B virus, drug-resistant mutations are likely to occur during antiviral drug treatment, and the drug resistance rate can reach 22% after two years of treatment. During long-term antiviral drug treatment, there is a lack of highly sensitive and Accurate detection methods enable drug-resistant strains to survive and reproduce in large numbers, resulting in treatment failure and considerable losses to the patient's body and economy....

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Abstract

The invention relates to a gene mutation detection primer and application, the primer includes an anchor sequence, a purine ring and a detection sequence, the anchor sequence at the is located at the primer 5'- end, forms more than 14 bases of complementary with a target sequence, the purine ring is formed as follows: 3-4 xanthosine, inosine or deoxidized inosine forms a bubble ring at the 5th-8th base position of the primer 3'- end, and the detection sequence is located at the primer 3'-end and is formed by 5-8 bases. According to the technical scheme of the primer, the bubble ring is inserted at the 5th-8th base position from the primer 3'- end, and the bubble ring is formed by '0' or '1' base counterpoint with a corresponding position of the target sequence, and the primer is given highly specific gene mutation detection ability, so that the primer can inhibit nonspecific amplification, improve the detection capability, and can be applied to the detection of low copy gene mutation detection, multiplex PCR (polymerase chain reaction), gene chip and other fields.

Description

technical field [0001] The invention relates to the technical field of detecting low-copy mutations, in particular to a primer for detecting gene mutations and its application. Background technique [0002] With the development of personalized medicine, SNP genotyping molecular diagnostic technology is playing an increasingly important role in the research of pharmacogenomics. Molecular diagnostic technology or genetic diagnostic technology has been gradually applied to clinical auxiliary diagnosis and medication . At present, my country's detection technology in SNP, mutant molecule and other diagnostic technologies is relatively backward. With the gradual implementation of the national 12th five-year development plan, especially the proposal of the clinical diagnosis technology development plan, the clinical diagnosis level in my country will be gradually improved. rapid development. It is of great practical significance to develop a rapid and accurate SNP detection metho...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6848C12Q2525/117
Inventor 王永忠
Owner 常州可尔生命科技有限公司
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