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Method for simultaneously detecting microsatellite locus stability and genome change on basis of second generation sequencing

A technology of microsatellite loci and genes, applied in the fields of genomics, biochemical equipment and methods, and microbial determination/inspection, etc. Effect

Active Publication Date: 2017-05-31
GUANGZHOU BURNING ROCK DX CO LTD
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Problems solved by technology

This method is the current gold standard detection method, but it requires additional detection samples, and requires the patient's normal tissue as a control for status determination, so it is not convenient from an operational point of view

Method used

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  • Method for simultaneously detecting microsatellite locus stability and genome change on basis of second generation sequencing
  • Method for simultaneously detecting microsatellite locus stability and genome change on basis of second generation sequencing
  • Method for simultaneously detecting microsatellite locus stability and genome change on basis of second generation sequencing

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Embodiment

[0049] The experimental samples involved in the examples are all from the domestic leading tertiary hospitals for the diagnosis and treatment of colorectal cancer, and all the experimental samples are clearly diagnosed as IHC-positive tumors and adjacent normal tissue samples. The instruments, reagents, kits and analysis software used in the experiments are all commercially available.

[0050] Method and steps

[0051] 1. Based on the next-generation sequencing method for gene combination (panel) detection, the specific steps are as follows:

[0052]DNA was extracted from tumor tissue and adjacent normal tissue using the QIAamp DNA FFPE tissue kit. Accurate quantification was performed with the dsDNA HS assay kits provided with the Qubit 3.0 Fluorometer. Then, the DNA was physically fragmented into 180-250bp fragments with an ultrasonic breaker CovarisM220, and then end repaired, phosphorylated, deoxyadenine was added to the 3' end, and the adapter was ligated. Then the DNA...

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Abstract

The invention relates to a method for simultaneously detecting microsatellite locus stability and genome change on the basis of second generation sequencing, especially an application of the detection method in assisting in diagnosing a colorectal cancer patient and a corresponding kit. According to the method provided by the invention, the sequence analysis for a plurality of samples can be completed at one time; the detection for genome change, such as, gene mutation and chromosome variation, is completed while the detection for the microsatellite state is completed; no additional experimental operation is required; the analysis time and the cost for detecting the patient are greatly saved; compared with PCR electrophoretic MSI detection gold standard, the detection method provided by the invention has the advantages of matching accuracy, high detection speed, lower price and no dependence on a matched normal sample, so that the detection mode is more convenient.

Description

technical field [0001] The present invention relates to a method for simultaneously detecting microsatellite stability and disease-related genes based on next-generation sequencing, especially the application of the detection method in auxiliary diagnosis, prognosis evaluation, or selection of treatment options for colorectal cancer patients. And the application of the corresponding test kit and detection reagent in the preparation of the test kit. Background technique [0002] Colorectal cancer (Colorectal Cancer, CRC) is the third highest incidence rate of various cancers in my country, accounting for the fifth cause of cancer death, its 5-year survival rate after radical resection is about 50%, postoperative recurrence and Metastasis is an important cause of its death. Patients with early colorectal cancer can usually be resected by surgery. Once metastasized, there are not many treatment options, and the five-year survival rate of patients is not ideal. Studies have fou...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68G06F19/22C12N15/11
CPCG16B30/00C12Q1/6869C12Q1/6886C12Q2600/118C12Q2600/156C12Q2535/122C12Q2600/112G16B20/20C12Q2600/158
Inventor 汉雨生张之宏揣少坤徐海燕刘成林
Owner GUANGZHOU BURNING ROCK DX CO LTD
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