Construction method and applications of tumor gene variation library for high-throughput sequencing detection

A technology of tumor gene and construction method, which is applied in the direction of chemical library, combinatorial chemistry, library creation, etc., to achieve the effect of low cost

Inactive Publication Date: 2017-11-03
XIAMEN SPACEGEN BIOTECH CO LTD
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  • Abstract
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Problems solved by technology

However, there is still no ideal tumor gene variation library fo

Method used

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  • Construction method and applications of tumor gene variation library for high-throughput sequencing detection
  • Construction method and applications of tumor gene variation library for high-throughput sequencing detection
  • Construction method and applications of tumor gene variation library for high-throughput sequencing detection

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Embodiment 1

[0088] A method for constructing a tumor gene variation library for high-throughput sequencing detection, covering human genes EGFR, K-ras, B-raf, PIK3CA, N-ras, c-MET, AKT1, HER2, c-KIT, PDGFRA A total of 389 somatic mutations on , ALK-EML4, ROS1 and RET, the information of the 389 somatic mutations is as follows:

[0089]

[0090]

[0091]

[0092]

[0093] Specifically include the following steps:

[0094] (1) Design the first basic amplification primer set for target genes EGFR, K-ras, B-raf, PIK3CA, N-ras, c-MET, AKT1, HER2, c-KIT and PDGFRA, the first basic amplification The 5' ends of the forward primer and the reverse primer of the primer set are provided with additional 2-5 Ts, and the first T near the 3' end of the 2-5 Ts has a PNA modification, and the first basic extension The Tm values ​​of the primer sets differ by no more than 1°C; specifically, the target gene EGFR is specifically exons 18, 19, 20 and 21 of the EGFR gene, and the target gene K-ras ...

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Abstract

The present invention discloses a construction method of a tumor gene variation library for high-throughput sequencing detection, wherein the tumor gene variation library covers 389 somatic cell mutations on human genes such as EGFR, K-Ras, B-raf, PIK3CA, N-ras, c-MET, AKT1, HER2, c-KIT, PDGFRA, ALK-EML4, ROS1 and RET. According to the present invention, the plurality of the target sequences are subjected to single-tube amplification to rapidly complete the construction of the library, wherein the construction time of the whole library is only 2-3 h, and the manual time is only 15-30 min; and by combining the obtained library and the high-throughput sequencing platform, the difficult problem that the multi-gene and multi-target detection of the somatic cells is required on the basis of the small amount of the clinical samples in tumors, genetic diseases and other diseases in the clinic can be effectively solved, and the cost is low.

Description

technical field [0001] The present invention specifically relates to a method for constructing a tumor gene variation library for high-throughput sequencing detection and its application. Background technique [0002] In 2015, China is expected to have 4.292 million new cancer cases and 2.814 million deaths. Lung cancer is the tumor with the highest incidence rate and the leading cause of cancer death. Gastric cancer, esophageal cancer and liver cancer are common tumors with higher morbidity and mortality in my country. [0003] Traditional methods of cancer treatment include surgery, chemotherapy, and radiotherapy. With the advancement of medical technology, targeted drug treatment of advanced lung cancer has increasingly become a new way, and the treatment varies according to the type of disease. Cancer patients with different driver gene types must be treated differently in treatment. For patients with targets, targeted therapy can be given priority. Targeted therapy...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C40B50/06
Inventor 陈琰郭飞飞
Owner XIAMEN SPACEGEN BIOTECH CO LTD
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