Primer pair, kit and method for methylation detection of lung cancer related gene SHOX2

A methylation and kit technology, applied in the field of primer pairs for lung cancer-related gene SHOX2 methylation detection, can solve the problems of cumbersome operation, low accuracy, low sensitivity, etc. Effectiveness of Diagnosis and Treatment

Inactive Publication Date: 2018-04-24
韩林志
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  • Claims
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Problems solved by technology

[0007] In order to solve the technical problems of cumbersome operation, low accuracy, and low sensitivity in the above-mentioned traditional methylation detection method, the present inv

Method used

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  • Primer pair, kit and method for methylation detection of lung cancer related gene SHOX2
  • Primer pair, kit and method for methylation detection of lung cancer related gene SHOX2
  • Primer pair, kit and method for methylation detection of lung cancer related gene SHOX2

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Embodiment 1: the preparation of kit

[0036] 1. Design and synthesis of primers and probes

[0037] For the promoter region of the SHOX2 gene in the human genome and the internal reference gene β-Actin (b-actin) (for the sequence, refer to the human whole genome sequence published in the NCBI database), use Primer Premier 3.0 and Methyl PrimerExpress v1.0 software to design a pair of Specific primers and probes.

[0038] Specific primers and probe sequences are shown in the table below:

[0039]

[0040] Note: Y is a degenerate base, that is, Y=C / T.

[0041] Among them, the 5' end of the first detection probe of SHOX2 is labeled with FAM as the luminescent group, the 5' end of the second detection probe of SHOX2 is labeled with VIC as the luminescent group, and the 5' end of the β-Actin detection probe is labeled with ROX as the luminescent group. group, and the 3' end uses MGB as a quenching group. The two SHOX2 probes jointly detect 6 CPG islands, increasing t...

Embodiment 2

[0068] Example 2: Method for detecting methylation of lung cancer using the above kit

[0069] 1. Technical principle

[0070] A pair of specific primers and probes for methylation detection were designed in the promoter regions of the SHOX2 gene and the internal reference gene β-Actin (b-actin) in the human genome. Then use the primers and probes to amplify the sample DNA converted by sulfite, and determine the methylation rate of the sample to be tested according to the relative fluorescence CT value of the PCR amplification results of SHOX2 and β-Actin genes. The rate indirectly judges the risk of lung cancer.

[0071] 2. Detection method

[0072] Step 1: Take the DNA extracted from the sample to be tested, carry out transformation treatment on it, and use the transformed DNA as a template for PCR;

[0073] Wherein, the reagents used in the conversion treatment are bisulfite or bisulfite, and other auxiliary materials (the corresponding kit is EpiTect Fast DNA BisuLfite ...

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Abstract

The invention relates to a primer pair for methylation detection of a lung cancer related gene SHOX2. The primer pair comprises an SHOX2 forward primer, an SHOX2 reverse primer, an SHOX2 detection probe, a beta-Actin forward primer, a beta-Actin reverse primer and a beta-Actin detection probe. The invention relates to a kit for the methylation detection of the lung cancer related gene SHOX2. The kit comprises PCR reaction liquid containing the primers and probes above, the PCR reaction liquid comprises the forward primers, the reverse primers, the detection probes, 10*PCR buffer, dNTP, nuclease-free water and Ex Taq enzymes. The invention further relates to a method for the methylation detection of the lung cancer related gene SHOX2. The kit and the detection method thereof have the advantages of being accurate in detection result, high in detection flux, good in specificity and sensitivity, short in detection time, convenient to use and capable of effectively meeting clinical requirements.

Description

technical field [0001] The invention relates to the technical field of in vitro diagnosis, in particular to a primer pair, a kit and a method for detecting methylation of a lung cancer related gene SHOX2. Background technique [0002] Lung cancer is the cancer with the highest incidence rate in China. Although histological biopsy and cytological detection can confirm the diagnosis of lung cancer, the disease is usually in a relatively advanced stage when it is detected, and the detection rate is not ideal. The methylation of the human dwarf homeobox gene SHOX2 is closely related to lung cancer. The detection of SHOX2 gene methylation provides a more effective option to solve this problem. A number of clinical trials have proved that its combined use with histological or cytological detection can improve the It is an effective tool to assist in the diagnosis of lung cancer. [0003] SHOX2 gene is a tumor suppressor gene closely related to lung cancer, and the methylation of ...

Claims

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Application Information

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IPC IPC(8): C12Q1/6886C12N15/11
CPCC12Q1/6886C12Q2600/154C12Q2600/166
Inventor 韩林志肖芳
Owner 韩林志
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