Cancer suppressor gene methylation high-throughput detection kit of colorectal cancer and application thereof

A detection kit and tumor suppressor gene technology, applied in the biological field, can solve problems such as low sensitivity, large sample demand, and inability to determine specific sequences, and achieve the effect of simple, fast and low cost

Active Publication Date: 2020-04-07
DALIAN GENTALKER BIO-TECH CO LTD
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  • Description
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  • Application Information

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Problems solved by technology

[0006] Before the popularization of sequencing technology, high-performance liquid chromatography (HPLC) was a classic method for methylation determination, but it could only measure the total DNA methylation level, and could not measure specific sequences.
The classic method for specific DNA sequence determination is Southern hybridization analysis, but this method has low sensitivity and requires a large amount of samples

Method used

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  • Cancer suppressor gene methylation high-throughput detection kit of colorectal cancer and application thereof
  • Cancer suppressor gene methylation high-throughput detection kit of colorectal cancer and application thereof
  • Cancer suppressor gene methylation high-throughput detection kit of colorectal cancer and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0037] According to the present invention, the nine colorectal cancer-related tumor suppressor gene promoter CpG island regions that have been obtained are used as methylation detection target regions, and DNA methylation mutation detection reagents for nine colorectal cancer-related tumor suppressor genes are designed. The kit, the kit firstly extracts the genomic DNA of the fecal sample through bisulfite treatment, and then undergoes methylation-specific multiplex PCR, digestion primers, fragment screening, the second round of enrichment amplification, mixed sequencing, analysis and other processes to obtain Sample test results.

[0038] 1. Sample collection, transportation and storage

[0039] The sample collection of the present invention selects a human feces sample, and collects 2g of the feces sample into a centrifuge tube filled with a preservation solution. Samples are transported in a normal temperature environment and delivered to the laboratory as soon as possible...

Embodiment 2

[0095] The present invention has collected a large number of clinical samples to verify the sensitivity performance of the detection method, collected 199 cases of clinical colorectal cancer / colorectal adenoma patient stool samples, and tested according to the experimental method described in Example 1, according to the obtained results The sensitivity verification data (as shown in Table 7) of the detection method of the present invention was obtained. Among the 199 clinical samples, 140 cases of stool samples from patients with stage I / II colorectal cancer and 59 cases of stool samples from patients with colorectal adenoma were formed. After the detection, the detection results obtained were 127 cases of positive detection results in 140 patients with stage I / II colorectal cancer, and the detection kit of the present invention can reach 91% for the detection sensitivity of stage I / II colorectal cancer; In 38 cases of rectal adenoma patients, the detection results were positiv...

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Abstract

The invention belongs to the technical field of biology and particularly relates to a cancer suppressor gene methylation high-throughput detection kit of the colorectal cancer and application thereof.The kit comprises methylation specific multiplex PCR amplification primers having the sequences shown as SEQ ID NO: 1-20 and label primers of which the sequences are shown as SEQ ID NO: 21-41. According to the invention, the target methylation detection areas of one or more genes can be captured at one time and the methylation mutation sequence information of one or more gene target regions of the sample is obtained, so that the methylation state of the target region can be accurately and effectively analyzed and thus the high-throughput, high-sensitivity and high-accuracy methylation detection on the target gene region becomes the reality. The method further has technical characteristics of low cost, simple and rapid operation and the like; and the sample library construction can be completed only through two rounds of PCR.

Description

technical field [0001] The invention belongs to the field of biotechnology, in particular to a high-throughput detection kit for colorectal cancer tumor suppressor gene methylation and its application. Background technique [0002] The incidence of colorectal cancer (CRC) ranks third among malignant tumors in the world, and it has become the third leading cause of new cancer cases and deaths. Regular screening can detect colorectal cancer early, allowing early intervention to prevent disease progression or even cure the disease. However, 60% to 70% of patients were diagnosed with advanced CRC at the time of first diagnosis. Statistics from the United States Preventive Services Task Force (USPSTF) show that if healthy people undergo routine regular inspections every year to achieve early diagnosis and treatment of CRC, about 60% of CRC patients can be prevented from dying, and the average five-year survival rate can reach From 46% to 73%. Therefore, an effective early scre...

Claims

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Application Information

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IPC IPC(8): C12Q1/6886C12Q1/6806C12Q1/6869
CPCC12Q1/6886C12Q1/6806C12Q1/6869C12Q2600/154C12Q2600/16C12Q2531/113C12Q2537/143C12Q2523/125C12Q2535/122Y02A50/30
Inventor 赵金银李宏志刘琦李杰
Owner DALIAN GENTALKER BIO-TECH CO LTD
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