High throughput sequencing method for detecting genes of multiple mutant types and applications thereof

A high-throughput, genetic technology, applied in the field of high-throughput sequencing, can solve problems such as limitations and high detection costs

Active Publication Date: 2018-08-21
合肥中科金臻生物医学有限公司 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0012] In summary, most of the gene detection methods in the prior art focus on the detection of point mutations, insertions/deletions, DNA copy number changes and other mutation types, while it is rare to combine multi-oncogene fusion problems with point mutations, insertions/deletions, DNA copy num...

Method used

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  • High throughput sequencing method for detecting genes of multiple mutant types and applications thereof
  • High throughput sequencing method for detecting genes of multiple mutant types and applications thereof
  • High throughput sequencing method for detecting genes of multiple mutant types and applications thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0096] The sample in this example is from a patient with stage IIB breast ductal carcinoma, and the source of the sample is human genomic DNA extracted from an isolated right breast tumor tissue sample.

[0097] Complete sample DNA extraction, hybridization and capture of target sequences, elution and enrichment and purification, library quality inspection, on-machine sequencing, and data processing and analysis according to the aforementioned 3-7 steps. The concentration of the obtained DNA library is 27.6ng / μL, and the results are as follows :

[0098] 8. Results

[0099] i) The detection information is as follows:

[0100] Table 4

[0101]

[0102]

[0103] ii) if figure 2 As shown, the L755_T759del mutation occurred in the ERBB2 gene of this sample, and the sequence data of the sample was compared with the paired normal human genome DNA. The identified mutation site is the L755_T759del (deletion of amino acid 755, 759) mutation in the ERBB2 gene. The mutation is...

Embodiment 2

[0110] The samples in this example are derived from human genomic DNA extracted from tumor tissue samples at the source site in vitro. The tumor was primary triple-negative breast cancer with axillary lymph node metastasis, and the biopsy results reported G3pT1cpN1 ductal type.

[0111] Complete sample DNA extraction, target sequence hybridization capture, elution and enrichment purification, library quality inspection, on-machine sequencing, and data processing and analysis according to the aforementioned 3 to 7 steps. The concentration of the obtained DNA library is 25.8ng / μL, and the results are as follows :

[0112] 8. Results

[0113] i) The detection information is as follows:

[0114] Table 6

[0115]

[0116] ii) if image 3 As shown, the p.T790M mutation occurred in the EGFR gene of the sample, and the sequence data of the sample was compared with the paired normal human genome DNA. The identified mutation site is the p.T790M (790 amino acid change) mutation i...

Embodiment 3

[0123] The sample in this example is derived from isolated left invasive ductal carcinoma of the breast, the pathological stage of the cancer is T2N0M0, and lymphovascular invasion.

[0124] Complete sample DNA extraction, target sequence hybridization capture, elution and enrichment purification, library quality inspection, on-machine sequencing, and data processing and analysis according to the aforementioned 3 to 7 steps. The concentration of the obtained DNA library is 26.2ng / μL, and the results are as follows :

[0125] 8. Results

[0126] i) The detection information is as follows:

[0127] Table 8

[0128]

[0129]

[0130] ii) if Figure 4 As shown, the ERBB2 gene of this sample has a copy number variation, and the sequence data of the sample is compared with the paired normal human genomic DNA. A copy number variation of >4 copies of the ERBB2 gene was found in the sample.

[0131] Table 9

[0132]

[0133] iii). Detailed analysis:

[0134] ERBB2 is a ...

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Abstract

The invention discloses a high throughput sequencing method for detecting genes of multiple mutant types and applications thereof, and belongs to the field of gene detection. By searching database, variation sites of genes for a cancer target treatment, chemotherapy drug resistant genes, and chemotherapy drug sensitive genes, 100 bp of downstream of the variation sites, and 100 bp of upstream of the variation sites are taken as a target section; a capture probe matched with the target section is synthesized, the target area of DNA of a sample is captured, a high throughput sequencing library is established; then high throughput sequencing is carried out to obtain the sequence of a target section; and the results of cancer related gene mutation, insertion mutation, delete mutation, mutationof copy number, and gene fusion mutation can be obtained. The provided method can detect multiple kinds of mutations of 105 cancer related genes at a time and has the characteristics of high capturing efficiency and good uniformity.

Description

technical field [0001] The invention belongs to the technical field of gene detection, and more specifically relates to a high-throughput sequencing method for detecting multiple mutation types of genes and its application, which can simultaneously detect multiple cancer-related gene fusions, copy number variations, point mutations and insertions Deletion variant. Background technique [0002] In China, the death rate caused by tumors is increasing year by year. Cancer has become an important factor affecting public health. In 2015, statistics showed that there were 4,292,000 new cancer patients and 2,814,000 deaths. With the popularization of diagnosis, the proportion of lung cancer diagnosis and death rate is increasing. Cancer rates are higher in rural areas than in urban areas. The tumor incidence rate in the United States is 45%, and the mortality rate is 30%, and this ratio is also increasing year by year. However, with the application of early tumor screening, di...

Claims

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Application Information

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IPC IPC(8): C12Q1/6858
CPCC12Q1/6858C12Q2531/113C12Q2535/122
Inventor 李河南王晓雪李浩浩李圆徐志兰花园林文楚洪波王伟
Owner 合肥中科金臻生物医学有限公司
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