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Kit for detecting CDKN2B gene mutation and quantitative detection method of CDKN2B gene mutation rate

A technology for gene variation and quantitative detection, applied in the direction of DNA/RNA fragments, recombinant DNA technology, biochemical equipment and methods, etc., can solve the problems of high cost, limited application, inconvenient use, etc., achieve high accuracy and save time and cost, detection high effect

Active Publication Date: 2017-09-15
苏州西山中科药物研究开发有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In the prior art, conventional quantitative PCR technology is used to detect the mutation rate of CDKN2B, which must be divided into two or more steps to obtain the mutation rate of the gene, which is inconvenient to use and expensive, which limits the application

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] Embodiment 1 Quantitative detection method of CDKN2B gene mutation rate

[0041] (1) Extract CDKN2B genomic DNA: Use DNeasy Blood&Tissue Kit (Qiagen product) to extract blood or tissue DNA according to the instructions;

[0042] (2) Quantitative PCR:

[0043] reaction system:

[0044] The PCR master mix is ​​2x Master mix ( Universal PCR Master Mix, ABI): 20ul

[0045] Primer (Primer)-MF (50nM): 0.5ul

[0046] Primer (Primer)-MR (50nM): 0.5ul

[0047] Probe-MP (250nM): 0.5ul

[0048] Hs03922281_s1: 1.0ul

[0049] Template DNA (10-20ng / ml): 10ul

[0050] Deionized water: 7.5ul

[0051] Total reaction volume = 40ul

[0052] The reaction conditions were: i), 50°C, 2 minutes; ii), 95°C, 10 minutes; iii), 95°C, 15 seconds; iv), 59°C, 1 minute; where iii) and iv) were performed for 45 cycles .

[0053] (3) Test results: coefficient of variation (%)=mutated CT value / non-mutated CT value×100.

[0054] When the coefficient of variation is more than 3 times greater t...

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Abstract

The invention provides a quantitative detection method of CDKN2B gene mutation rate, specifically, two indicators of expression level and mutation rate of CDKN2B are detected in a reaction system for quantitative detection of the CDKN2B gene mutation rate, and compared with conventional quantitative PCR technique which must comprises two steps or more steps to obtain the gene mutation rate in the prior art, the quantitative detection method has the advantages of more convenient operation, saving time, lower cost and more comparable data.

Description

technical field [0001] The invention relates to the field of gene mutation rate detection methods, and more particularly, the invention relates to a kit for detecting CDKN2B gene mutation and a quantitative detection method for CDKN2B gene mutation rate. Background technique [0002] CDKN2B (Cyclin-dependent kinase 4inhibitor B) is a cell cycle kinase inhibitor, adjacent to the tumor suppressor gene CDKN2A, a region of frequent mutations that can be detected in many tumor cases. The gene encodes a cell cycle kinase inhibitor, which forms a complex with CDK4 or CDK6 to inhibit the activity of cell cycle kinases. By controlling the process of cell cycle G1, the functional protein is a cell growth regulator. The expression of this gene was induced by TGFbeta, suggesting involvement in the growth regulation of TGFbeta. [0003] At present, the detection of CDKN2B gene is mainly used for clinical prediction of diabetic patients, and since many anti-tumor drugs based on CDK have ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6883C12Q2600/156
Inventor 张海鹰严杰董文明金嘉敏
Owner 苏州西山中科药物研究开发有限公司