Method for carrying out high-flux sequencing on physiological status of individual to which cfDNA (Circulating cell-free DNA) belongs based on blood free DNA, and application thereof

A technology for individual physiology and sequencing analysis, applied in the field of biomedicine, can solve problems such as false positives, complex bioinformatics predictions, and ctDNA abundance failure, and achieve the effects of avoiding human errors, simplifying processing procedures, and avoiding the introduction of errors

Inactive Publication Date: 2019-04-26
SOUTHEAST UNIV
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Problems solved by technology

However, except for some advanced tumors that release a large amount of ctDNA, the abundance of ctDNA in most tumor patients does not meet this standard
This makes detecting ctDNA very difficult, especially in the early stages of cancer
At present, the sensitivity and accuracy of ctDNA detection are mainly improved by increasing the sequencing depth, but increasing the sequencing depth may bring false positive results, because non-tumor-derived DNA may also carry various tumor-related

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  • Method for carrying out high-flux sequencing on physiological status of individual to which cfDNA (Circulating cell-free DNA) belongs based on blood free DNA, and application thereof
  • Method for carrying out high-flux sequencing on physiological status of individual to which cfDNA (Circulating cell-free DNA) belongs based on blood free DNA, and application thereof
  • Method for carrying out high-flux sequencing on physiological status of individual to which cfDNA (Circulating cell-free DNA) belongs based on blood free DNA, and application thereof

Examples

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Effect test

Embodiment 1

[0040] Example 1 Esophageal cancer detection based on cfDNA NGS

[0041] Experimental materials and methods

[0042] Sample collection: A total of 19 whole blood samples were collected with the help of Jinling Hospital Affiliated to Nanjing University School of Medicine (Nanjing, China). Among them, four were collected from normal subjects, eleven were collected from preoperative esophageal cancer patients, and five were collected from postoperative esophageal cancer patients (Table 1).

[0043] Table 1. cfDNA source information

[0044]

[0045]

[0046] Cell-free DNA extraction: Whole blood samples were centrifuged at 1,600g for 15 minutes at 4°C, and the supernatant was transferred to a new centrifuge tube. The tube containing the supernatant was centrifuged at 16,000g for 10 minutes at 4°C, and the supernatant was the plasma. All plasma was stored at -80°C for later use. Using the Plasma Circulating DNA kit (TIANGEN, DP339), 200 μL plasma was used as the starting...

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Abstract

The invention discloses a method for carrying out high-flux sequencing on the physiological status of an individual to which cfDNA (Circulating cell-free DNA) belongs based on blood free DNA, and application thereof. The method comprises the following steps: separating the blood free DNA; constructing the blood free DNA to obtain a high-flux sequencing library; applying DNA high-flux sequencing tocarry out sequencing on a blood free DNA library; carrying out biological information analysis on reads obtained by sequencing; on the basis of a biological information analysis result, judging the physiological status of an individual to which cfDNA to be detected belongs. According to the method, a cfDNA detection and analysis process is simplified, and the cfDNA does not need to be subjected to preprocessing, including fragment selection, methylation enrichment and the like, before sequencing; after sequencing is carried out, complex fragment selection, terminal coordinate discovery, bioinformatics modeling and the like are not required after sequencing, and the judgment of the physiological status of an individual to which the cfDNA belongs can be realized. The method can be applied to auxiliary cancer non-invasive detection or the preparation of an auxiliary cancer non-invasive detection kit.

Description

technical field [0001] The invention belongs to the field of biomedical technology, and specifically relates to a method for analyzing the physiological state of individuals to which cfDNA belongs based on high-throughput sequencing of free blood DNA and its application in non-invasive detection of cancer and preparation of auxiliary non-invasive detection kits for cancer. Background technique [0002] Since the birth of next-generation sequencing (NGS) technology, it has become an important research, detection and diagnosis technology, and has had a profound impact on the research of life science, biomedicine and medicine. NGS technology has been widely used in the field of non-invasive detection of cancer, and plays an important role in cancer diagnosis, typing, mutation discovery, medication guidance, treatment monitoring, and prognosis judgment. [0003] Tumor cells release DNA fragments into the blood circulation, and this source of circulating cell-free DNA (cfDNA) is ...

Claims

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Application Information

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IPC IPC(8): C12Q1/6869C12Q1/6886
CPCC12Q1/6869C12Q1/6886C12Q2525/191C12Q2531/113C12Q2535/122
Inventor 王进科武剑刘世财
Owner SOUTHEAST UNIV
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