Kit and method for detecting HER2 gene amplification

A gene amplification and detection method technology, applied in the biological field, can solve problems such as low accuracy, achieve the effects of good accuracy, improved detection specificity and accuracy, and high precision

Pending Publication Date: 2020-10-30
苏州索真生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] The purpose of the present invention is to provide a kit and method for detecting HER2 gene amplification based on digital microdroplet PC

Method used

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  • Kit and method for detecting HER2 gene amplification
  • Kit and method for detecting HER2 gene amplification
  • Kit and method for detecting HER2 gene amplification

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] Embodiment 1 detects the kit of HER2 gene amplification

[0035] 1. Design and obtain HER2, EIF2C1, TFF3, and CEP17 genes, according to the above-mentioned gene sequences published in the existing gene database.

[0036] 2. Multiplex Digital Droplet PCR Primer Probe Design

[0037] Use primer5 to design primers and probe sets to quantify the DNA copy numbers of HER2, EIF2C1, TFF3, and CEP17, and select target sequences in conserved regions that are different from common single nucleotide polymorphisms or copy number variations. Use oligo software to screen potential secondary structures, dimers, hairpin structures, etc. Base recombination optimization was performed to minimize unwanted primer-primer interactions in multiplex PCR reactions.

[0038] The designed primer sequences are as follows:

[0039]

[0040] 3. Synthesize the primers and probes designed above.

Embodiment 2

[0041] Example 2 Using the kit for detecting abnormal HER2 gene amplification of the present invention to detect abnormal HER2 gene amplification of samples

[0042] 1. Formulation of sampling plan and extraction of cell-free DNA

[0043] Sample collection and preparation All experiments were performed in accordance with the Declaration of Helsinki and national guidelines. A total of 60 plasma samples and 60X10-15 pathological white slices (per person) were collected from breast cancer patients. The tumor DNA was isolated from 5 pathological slides using the QIAgen_FFPE_DNA_kit, cfDNA was eluted with 30 μl deionized water, and stored at -80°C. Plasma samples were isolated from free DNA (cfDNA) from 2 ml of plasma using a magmax cfDNA extraction kit (Thermo Fisher Scientific, MA), eluted with 30 μl of deionized water, and stored at -80°C. Using Covaris M220 sonicator (ThermoFisher Scientific, MA), the DNA extracted from tumor tissue was sheared into fragments of about 200 bp,...

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Abstract

The invention aims to provide a kit and method for detecting HER2 gene amplification based on a digital droplet PCR technology so as to solve the technical problem of low accuracy of an HER2 gene copynumber variation judgment method in the prior art.

Description

technical field [0001] The invention relates to the field of biotechnology, and provides a kit and a method for detecting HER2 gene amplification based on a digital droplet PCR method. Background technique [0002] Breast cancer is a malignant tumor that seriously threatens women's health, and its incidence is increasing year by year. 25%-30% of breast cancers have HER2 gene amplification and protein positive expression. Studies have confirmed that these patients have shorter OS and DFS. Herceptin, a monoclonal antibody drug targeting HER2 Zhuzumab) has been widely used clinically, and is of great significance to the treatment of breast cancer. [0003] HER2, also known as erbB2, is a member of the epidermal growth factor (EGFR) family. It is located on chromosome 17q12-21.32 and encodes a 185KDa cell receptor. It plays an important role in tumor proliferation, metastasis and invasion. Patients with HER2-positive tumors have The response to conventional chemotherapy and ta...

Claims

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Application Information

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IPC IPC(8): C12Q1/6886C12Q1/6851C12N15/11
CPCC12Q1/6886C12Q1/6851C12Q2600/158C12Q2600/118C12Q2600/166C12Q2600/16C12Q2531/113C12Q2537/165C12Q2545/101C12Q2563/107C12Q2563/159C12Q2537/143C12Q2537/16
Inventor 陈谦刘弘扬刘泓
Owner 苏州索真生物技术有限公司
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