High-flux detection method of circulating DNA liver cancer driver gene

A gene-driven, high-throughput technology, applied in the fields of biochemical equipment and methods, microbial determination/examination, etc., can solve the problems of patient pain, invasiveness, and difficulty in obtaining patient samples, and achieve strong predictability and accuracy. high effect

Pending Publication Date: 2018-02-23
JIAXING YUNYING MEDICAL INSPECTION CO LTD
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Problems solved by technology

Based on the detection of tumor tissue samples, there are two problems: 1. Most of the tumor tissue comes from biopsy techniques, namely puncture and biopsy forceps biopsy, which will bring great pain to the patient and is an invasive detection; 2. For some advanced For patients who cannot undergo puncture or surgery, it is difficult to obtain patient samples, so it is difficult to carry out targeted drugs for patients based on the test results
Therefore, a problem arises that more treatment options require more samples from patients

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  • High-flux detection method of circulating DNA liver cancer driver gene

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Embodiment Construction

[0044] A detection method of circulating tumor DNA liver cancer gene of the present invention, comprises the steps:

[0045] 1. Sample library preparation:

[0046] (1) Pipette 60 ul of purified DNA (cfDNA) and 40 ul of Endprep mix into a single PCR tube, pipette to mix, and briefly centrifuge for end repair. The reaction conditions are 30°C and 30 min.

[0047] (2) Take out the magnetic beads half an hour in advance to return to room temperature, shake and mix, take 180ul of magnetic beads and add them to the end-repaired PCR product, beat and mix, and incubate in the greenhouse for 5 minutes.

[0048] (3) Put the PCR single tube on the magnetic stand, let it stand for 5 minutes, remove the supernatant, keep the PCR tube on the magnetic stand, add 200ul of 80% ethanol (newly prepared) to rinse, let stand for 30s, remove the supernatant, 80 Rinse twice with % ethanol, and dry at room temperature for 10 minutes (with 2 minutes remaining, use a small grab to suck up the residue...

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Abstract

The invention discloses a high-flux detection method of a circulating DNA liver cancer driver gene. According to the method, a mutant gene which is the most common of the existing liver cancer is selected and combined with a second-generation high-flux sequencing technology to more comprehensively detect a gene mutant condition of the liver cancer. Since a blood plasma sample of a patient is adopted, the method can greatly reduce the detection injury of a patient; secondly, the whole-exome detection is performed on 75 liver cancer genes, the method is more comprehensive compared to the detection method based on real-time PCR and first-generation Sanger sequencing technology; and since only 75 genes are detected, the sample can be detected in a greater sequencing depth, so that the detection precision can reach 0.1 percent. Therefore, a reference can be provided for more and better individual precision therapeutic schemes of the patient.

Description

technical field [0001] The invention relates to a gene mutation detection method, in particular to a high-throughput detection method of circulating tumor DNA liver cancer driver genes. Background technique [0002] Hepatocellular carcinoma is the fifth most common cancer and ranks third in cancer-related mortality worldwide. China is a big country with liver cancer. According to national cancer statistics in 2012, the incidence of liver cancer in Chinese men was 293,318, and the incidence of liver cancer in women was 101,452. A total of 394,770 men and women were diagnosed, accounting for 52.7% of new liver cancer patients in the world. . It is estimated that in 2016, the incidence of liver cancer in Chinese men is 326,698, and the incidence of liver cancer in women is 111,017. The sum of the two is 437,715, which is equivalent to the population of a county in China. The form of cancer prevention and treatment is very severe. How to screen effective molecular markers for ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6869
CPCC12Q1/6869C12Q2535/122C12Q2565/519C12Q2531/113
Inventor 张道允巩子英叶建伟王伟
Owner JIAXING YUNYING MEDICAL INSPECTION CO LTD
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