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Multiple PCR primers for detecting non-small cell lung cancer oncogene mutation based on high-throughput sequencing, kit and method

A non-small cell lung cancer, high-throughput technology, applied in the field of multiplex PCR primers, can solve the problems of inability to meet the needs of non-small cell lung cancer detection of gene mutations, difficult to obtain materials for advanced patients, false positives and false negatives, etc., and achieve a simple detection method. The effect of easy operation, fast detection speed and high accuracy

Active Publication Date: 2018-02-23
GUANGZHOU FOREVERGEN HEALTH TECH CO LTD +1
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AI Technical Summary

Problems solved by technology

[0007] At present, the detection of non-small cell lung cancer in the market is mainly at the detection level of a single or a few sites. The detection methods generally use fluorescent PCR, chip analysis, etc. The common disadvantage of these methods is that only one mutation or a segment can be detected at a time. Gene mutations, low detection throughput, difficult to interpret results, poor repeatability, many false positives and false negatives, etc.
At the same time, due to the difficulty in obtaining materials for advanced patients and the small sample size, it often cannot meet the needs of the above-mentioned method for detecting gene mutations in non-small cell lung cancer.
Although some existing deep sequencing methods based on high-throughput sequencing technology can meet clinical needs in terms of throughput, they cannot be popularized in clinical applications due to high prices.

Method used

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  • Multiple PCR primers for detecting non-small cell lung cancer oncogene mutation based on high-throughput sequencing, kit and method

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Embodiment 1

[0032] The samples are: Genomic DNA of normal blood, cfDNA of normal human plasma, MCF10A cell line genomic DNA, liver cancer FFPE genomic DNA, using DNA extraction kits to extract genomic DNA from various types of samples, using SEQ No.1~SEQ No.392 performs multiple PCR amplification on the extracted genomic DNA, and then uses the IlluminaNextSeq 500 sequencer for sequencing analysis. The specific implementation method is as follows:

[0033] A. Genomic DNA extraction: use Qiagen DNeasy Blood&Tissue Kit, and refer to the kit instructions to extract genomic DNA. The obtained genomic DNA is subjected to gel electrophoresis and quality detection using NanoDrop 100. It is required that the genomic DNA is not significantly degraded, the concentration is greater than 20ng / μL, A260 / A280>1.8, A260 / A230>2.0, and Qubit 3.0 is used for accurate quantification.

[0034] B. The first round of multiplex PCR amplification: use the amplification primer mixture R1-barcode (SEQ ID NO:1~SEQ IDN...

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Abstract

The invention provides multiple PCR primers for detecting non-small cell lung cancer oncogene mutation based on high-throughput sequencing, a kit and a method. The method provided by the invention isan efficient and reliable high-throughput sequencing sequence enriching method. According to the detection method on the basis of PCR combined with high-throughput sequencing technique provided by theinvention, the specific mutation site of the related gene can be directly responded and the method can be directly used for guiding the clinic non-small cell lung cancer targeted drug and early diagnosing cancer or assisting in diagnosing and screening the cancer and monitoring after the cancer healing.

Description

technical field [0001] The invention relates to the detection field, in particular to a multiplex PCR primer and a method for detecting non-small cell lung cancer oncogene mutations based on high-throughput sequencing. Background technique [0002] The morbidity and mortality of lung cancer rank first among tumors. In the past three decades, the mortality rate of lung cancer in my country has increased by 465%, surpassing liver cancer to become the cancer with the highest mortality rate. [0003] Lung cancer can be divided into small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC) according to histology. Non-small cell lung cancer accounts for about 80% of the total lung cancer. It is mainly divided into Adenocarcinoma (~50%) and squamous cell carcinoma (~40%) are two subtypes. [0004] The traditional treatment method is to select a chemotherapy regimen based on the patient's clinical characteristics and pathological types, but the progression-free period (...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6869C12N15/11
CPCC12Q1/6869C12Q2537/143C12Q2535/122C12Q2563/185
Inventor 林钊张燕菲张纪斌何广良周艳河何铭辉
Owner GUANGZHOU FOREVERGEN HEALTH TECH CO LTD
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