DNA methylation kit for colorectal cancer detection and detection method thereof

A colorectal cancer and methylation technology, which is applied in the field of determining the methylation status of one or more target gene targets, can solve the problems of low specificity and sensitivity of detection methods, and improve the positive detection rate , the effect of improving specificity

Pending Publication Date: 2020-04-07
深圳市新合生物医疗科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0015] The purpose of the present invention is to provide a blood polygene joint detection primer probe combination kit for early diagn...

Method used

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  • DNA methylation kit for colorectal cancer detection and detection method thereof
  • DNA methylation kit for colorectal cancer detection and detection method thereof
  • DNA methylation kit for colorectal cancer detection and detection method thereof

Examples

Experimental program
Comparison scheme
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Embodiment 1

[0061] Example 1: Multiplex qPCR detection of colorectal cancer methylation on healthy human cfDNA samples using the kit of the present invention

[0062] (1) Experimental materials

[0063] 1. Healthy human plasma (purchased from Bloodworks NW);

[0064] 2. QIAamp Circulatory System Nucleic Acid Extraction Kit (purchased from Qiagen);

[0065] 3. EZ DNA Methylation-Lightning Kit (purchased from Zymo Research);

[0066] 4. The kit of the present invention (which includes primers and probe combination 1 for detecting target SEPT9 and primer and probe combination 1 for MLH1, ACTB amplification primers and probes);

[0067] 5. AmpliTaq Gold TM DNA polymerase and buffer reagents (purchased from Thermo Fisher).

[0068] (2) Experimental method

[0069] 1. DNA extraction from plasma

[0070] cfDNA was extracted from 5 healthy human plasma samples using the QIAamp Circulatory System Nucleic Acid Extraction Kit, and the steps were performed according to the kit instructions.

...

Embodiment 2

[0083] Example 2: Multiplex qPCR detection of colorectal cancer methylation on ctDNA samples of colorectal cancer patients using the kit of the present invention

[0084] (1) Experimental materials

[0085] 1. Plasma from patients with colorectal cancer (purchased from Bloodworks NW);

[0086] 2. QIAamp Circulatory System Nucleic Acid Extraction Kit (purchased from Qiagen);

[0087] 3. EZ DNA Methylation-Lightning Kit (purchased from Zymo Research);

[0088] 4. The kit of the present invention (which includes primers and probe combination 1 for detecting target SEPT9 and primer and probe combination 1 for MLH1, ACTB amplification primers and probes);

[0089] 5. AmpliTaq Gold TM DNA polymerase and buffer reagents (purchased from Thermo Fisher).

[0090] (2) Experimental method

[0091] 1. DNA extraction from plasma

[0092] ctDNA was extracted from 5 plasma samples of patients with colorectal cancer using the QIAamp Circulatory System Nucleic Acid Extraction Kit, and th...

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Abstract

The invention discloses a DNA methylation kit for colorectal cancer detection and a detection method thereof. The kit comprises a specific primer and a probe which are used for detecting or measuringa methylation state or level of one or more specific genes in DNA of a tested sample, wherein the specific genes are SEPT9, MLH1 and ACTB. The kit provided by the invention can significantly improve the positive detection rate and specificity (real negative rate) of early colorectal cancer; the detection sensitivity of a biomarker can be improved to a pg/nanogram-grade DNA molecule, and the improvement of the detection sensitivity is realized by optimizing a specific nucleotide sequence primer and a DNA probe and improving a bisulfite treatment method of plasma free DNA.

Description

technical field [0001] The invention belongs to the technical field of biotechnology and DNA detection, and in particular relates to a DNA methylation qPCR kit for colorectal cancer detection and its application method, in particular to a method for methylation qPCR using plasma free DNA A kit for detecting or screening colorectal cancer by determining the methylation status of one or more target gene targets (such as SEPT9 and MLH1) and a method of use thereof. Background technique [0002] Colorectal cancer (CRC) is the third most common cancer worldwide, killing 600,000 people each year. For CRC, early recognition and early intervention are the best ways to reduce mortality. The current screening method is colonoscopy with fecal occult blood and immunohistochemical testing (gFOBT / FIT). These screening methods are underutilized due to the discomfort of colonoscopy and the distaste for bowel products. Therefore, assessment of CRC risk using reliable non-invasive blood-ba...

Claims

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Application Information

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IPC IPC(8): C12Q1/6886C12Q1/6851
CPCC12Q1/6886C12Q1/6851C12Q2600/154C12Q2600/16C12Q2531/113C12Q2561/101C12Q2537/143C12Q2523/125
Inventor 王志强万季潘晓新夏迪余涛金泰庆罗海燕蔡雪儿宋麒
Owner 深圳市新合生物医疗科技有限公司
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