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Method and device for detecting copy number variation based on amplicon next generation sequencing

A technology of copy number variation and next-generation sequencing, which is applied in the field of copy number variation detection based on amplicon-based next-generation sequencing, to achieve the effect of preventing false positive detection and increasing sensitivity

Active Publication Date: 2017-02-01
天津诺禾致源生物信息科技有限公司
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  • Claims
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Problems solved by technology

However, the accuracy of the detection effect of this method needs to be further improved.

Method used

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  • Method and device for detecting copy number variation based on amplicon next generation sequencing
  • Method and device for detecting copy number variation based on amplicon next generation sequencing
  • Method and device for detecting copy number variation based on amplicon next generation sequencing

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Embodiment 1

[0027] In this embodiment, the sample to be tested is the SK-BR-3 cell line (purchased from the American Type Culture Collection Bank), which is a Her2 amplification-positive cell line, and has a 9-fold positive amplification of the HER2 gene after 3d-pcr testing. increase.

[0028] The specific steps of this embodiment are as follows (all reagents are purchased from Thermo Fisher in this example):

[0029] 1) Extract DNA from the sample to be tested, quantify it with a fluorometer (Qubit), concentrate or dilute with nuclease-free water to make the concentration 5ng / ul and the volume 6ul.

[0030] 2) Multiplex PCR technology was used to amplify the target region, and the PCR reaction system is shown in Table 1.

[0031] Table 1

[0032] Reagent volume High Fidelity Multiplex Amplification Reaction Mix 4μl primer mix 10μl plasma cell-free DNA 6μl

[0033] Among them, the specific sequences of the primers are shown in Table 2.

[0034] Tabl...

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Abstract

The invention discloses a method and device for detecting copy number variation based on amplicon next generation sequencing. The method comprises the following steps: S1, extracting DNAs (Deoxyribonucleic Acid) of a sample to be detected and a negative comparison sample respectively and amplifying a target region with relatively mutated target characters through a plurality of amplicons, wherein the negative comparison sample is a sample with target character related genes which are not mutated; S2, carrying out next generation sequencing to obtain a sequence of the target region; S3, comparing the sequence of the target region and a reference genome, and carrying out data information processing to obtain a coverage degree of each amplicon; and S4, establishing a normal distribution model of the negative comparison sample on the coverage degree of each amplicon respectively and taking the normal distribution model as background noises, carrying out bilateral checking on the coverage degree of each amplicon of the sample to be detected and the normal distribution model, and taking amplicons which do not belong to normal distribution as copy number variation and positively detecting. By adopting the technical scheme provided by the invention, the accuracy of copy number variation detection based on the amplicon next generation sequencing is improved.

Description

technical field [0001] The present invention relates to the field of biology, in particular, to a method and device for detecting copy number variation based on amplicon next-generation sequencing. Background technique [0002] Multiple amplicon next-generation sequencing is to customize the genomic region of interest into specific amplification primers and genomic DNA for specific amplification, enrich the DNA fragments of the target genomic region, and then use the second-generation sequencing technology for sequencing research strategy. Multiple amplicon next-generation sequencing is a hot technology in genomics research, mainly because it consumes a small amount of cost and time. At the same cost, researchers can study more samples and measure deeper depths. As a powerful and effective technology, it plays a unique role in next-generation high-throughput sequencing, and its application fields are becoming more and more extensive. [0003] Ion AmpliSeq from Thermo Fish...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G06F19/20
CPCG16B25/00
Inventor 朱嘉麒王棪张振宇马丽娟
Owner 天津诺禾致源生物信息科技有限公司
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