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Genetic marker combination and application thereof

A gene marker and gene technology, applied in the field of biomedicine, can solve the problems such as the difficulty of gene methylation markers, the overlapping of colorectal cancer detection value and performance, and the limited sensitivity of detection.

Active Publication Date: 2019-10-11
CREATIVE BIOSCIENCES (GUANGZHOU) CO LTD
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  • Claims
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AI Technical Summary

Problems solved by technology

The existing method for detecting a single methylation marker of colorectal cancer has the following two limitations: First, the existing research and product detection methods all detect one marker in one PCR reaction well, and the detection method is not able to Enables multiplex detection of methylated markers in one PCR reaction well
Second, when existing methylation markers for colorectal cancer detection such as SDC2, ITAG4, and Septin 9 detect colorectal cancer by detecting the methylation level of a single gene, the detection sensitivity is limited by genetic factors
The specificity of these individual markers for the detection of colorectal cancer can reach more than 90%, but the sensitivity when the specificity reaches more than 90% cannot reach more than 90%.
[0007] Although a single methylation marker of colorectal cancer can detect part of colorectal cancer, it is necessary to combine colorectal cancer-related markers to detect as many colorectal cancer patients as possible through molecular biology methods. The problem is: 1. The detection value of most colorectal cancer gene methylation markers for colorectal cancer is overlapping, that is, the detection status of the same colorectal cancer patient sample is consistent among most methylation markers of
Therefore, it is very difficult to screen out gene methylation markers that can complement each other in the detection of colorectal cancer, requiring a lot of research work
In addition, the more gene methylation markers, the false positive phenomenon of the detection system will also be superimposed, and the specificity of detection will decrease

Method used

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  • Genetic marker combination and application thereof
  • Genetic marker combination and application thereof
  • Genetic marker combination and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0127] Select 935 stool samples (359 cases of colorectal cancer, 67 cases of adenoma (≧1cm), 509 cases of non-tumor individuals, all confirmed by colonoscopy or pathology), grind and centrifuge, add 50ul capture magnetic beads (containing SDC2, COL4A2 , ITGA4 and the capture sequence of the internal reference gene ACTB), and operate according to the technical scheme as described below,

[0128] The technical solution is as follows:

[0129] 1) Collect stool samples from normal people and colorectal tumor patients with colonoscopy pathological results, mix and grind 1g of stool: 4mL of protective solution, centrifuge at 5000rpm for 10min, take the supernatant and discard the sediment;

[0130] 2) Take out 10mL of supernatant and centrifuge again, take 3.2mL of supernatant, add 2mL of lysate and 50ul of magnetic capture beads M1, incubate at 95°C for 15min, then place at room temperature for 30min;

[0131] 3) Discard part of the supernatant on the magnetic stand, wash the magn...

Embodiment 2

[0173] Select 23 patients diagnosed with colorectal cancer, collect the stool samples of the patients before operation and 3-6 months after operation, and use the marker combination of the present invention (SDC2, COL4A1 / COL4A2 and ITGA4) to detect the preoperative and postoperative A total of 46 stool samples were collected, and the methylation levels of the marker combination in the stool before and after surgery were compared.

[0174] The detection process and result judgment criteria of the marker combination are the same as in Example 1. Target gene CT value ≤ 38 was judged as positive, and CT value > 38 was judged as negative. The result is as figure 2 shown.

[0175] The results showed that the preoperative test results of these 23 patients with colorectal cancer were all positive, and the stool samples were tested again after the operation, and the results were all negative. It shows that after tumor resection, the methylation level of this marker combination is s...

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Abstract

The invention relates to a genetic marker combination and application thereof, belongs to the field of biomedicine, and particularly relates to a tumor marker, a methylation detection reagent, a kit and application thereof. Through studies, it is found that by detecting the methylation level of an SDC2, COL4A1 / COL4A2 and ITGA4 gene combination promoter region, colorectal cancer specimens can be distinguished from fecal specimens well. A methylation reagent methylation detection reagent containing the gene combination is utilized to detect colorectal cancer, and the detection sensitivity and specificity for the intestinal cancer are very high, and can reach 90% or above.

Description

technical field [0001] The invention belongs to the field of biomedicine, in particular to tumor markers, methylation detection reagents, kits and applications thereof. [0002] Background of the invention [0003] Colorectal cancer, also known as colorectal cancer, is a common malignant tumor of the digestive tract. Its incidence rate is increasing year by year in my country. In some coastal areas of my country such as Shanghai and Guangzhou, the incidence rate of colorectal cancer has jumped to the second place, second only to lung cancer. It is currently believed that the formation of colorectal cancer is the result of the accumulation of genetic and epigenetic defects. The onset of colorectal cancer is hidden in the early stage, and there are often no obvious symptoms. Symptoms such as blood in the stool, abdominal pain, and diarrhea may appear in the late stage. And when symptoms appear, it is often late, which brings great misery and expensive treatment costs to patie...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6886C12N15/11
CPCC12Q1/6886C12Q2600/154C12Q1/6827C12Q2537/143C12Q2537/164C12Q2545/10C12Q1/6816G16B40/10
Inventor 吴孝林刘相林罗茵邹鸿志
Owner CREATIVE BIOSCIENCES (GUANGZHOU) CO LTD
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