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Detection kit and detection method for 19 deletion mutations of EGFR gene exon 19

A detection kit and deletion mutation technology, which is applied in the fields of biotechnology and medicine, can solve problems such as unsatisfactory detection results, and achieve simple and objective result interpretation, high detection sensitivity, and simple and effective detection methods

Active Publication Date: 2013-11-06
WUHAN YZY MEDICAL SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0010] The purpose of the embodiments of the present invention is to provide a detection kit for 19 deletion mutations of exon 19 of the EGFR gene and its detection method, aiming to solve the problem of unsatisfactory detection results for such deletion mutations in the prior art

Method used

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  • Detection kit and detection method for 19 deletion mutations of EGFR gene exon 19
  • Detection kit and detection method for 19 deletion mutations of EGFR gene exon 19
  • Detection kit and detection method for 19 deletion mutations of EGFR gene exon 19

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0053] Preparation of the EGFR gene exon 19 mutation detection kit specifically includes the following steps:

[0054] 1. Synthetic primers and probe sequences

[0055] Synthetic primer sequence SEQ ID NO:1, SEQ ID NO:2; Synthetic specific probe sequence SEQ ID NO:3, and labeled with FAM fluorescent group at the 5' end, and labeled with quenching groups NFQ and MGB at the 3' end modifying group.

[0056] The above primer sequences were respectively prepared into 100 μM mother solution for storage, and the above probe sequences were respectively prepared into 100 μM mother solution for storage.

[0057] 2. Synthetic Blocking Nucleic Acid Sequence

[0058] Synthetic blocking nucleic acid sequence SEQ ID NO:4, and a phosphate group (PO) at the 3' end 4 ) modification.

[0059] The blocking nucleic acid was prepared as a 100 μM stock solution for storage.

[0060] 3. Preparation of fluorescent quantitative PCR reaction system

[0061] Prepare the quality control detection re...

Embodiment 2

[0067] The EGFR gene exon 19 mutation detection kit prepared in Example 1 was used to detect the samples to be tested.

[0068] In this embodiment, 41 cases of paraffin-embedded tissue sections of patients with clinical and pathological diagnosis of non-small cell lung cancer were collected, and genomic DNA was extracted therefrom, and the samples to be tested were detected with the EGFR gene exon 19 mutation detection kit obtained in Example 1 Whether there is a deletion mutation of exon 19 of the EGFR gene in the test, and the traditional sequencing method is used to verify. The specific operation steps are:

[0069] 1. Extraction of Genomic DNA from Samples

[0070]Genomic DNA was extracted from the tissue samples of the lung cancer patients above using a DNA extraction kit (QIAamp DNA FFPE Tissue Kit, Cat No. 56404). The DNA extraction method is carried out according to the instructions, and the operation steps are briefly described as follows: Place the clinically collec...

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Abstract

Being applicable to the fields of biotechnologies and medical science, the invention relates to a detection kit and a detection method for 19 deletion mutations of EGFR (epidermal growth factor receptor) gene exon 19. The detection kit provided in the invention includes a first reagent and a second reagent, wherein the first reagent contains the primer pair SEQ ID NO:1 and SEQ ID NO:2, the probe SEQ ID NO:3, and the second reagent contains the primer pair SEQ ID NO:1 and SEQ ID NO:2, the probe SEQ ID NO:3 and the block nucleotide sequence SEQ ID NO:4. Specifically, the 5' terminal of the probe SEQ ID NO:3 is in connection with a fluorescence group, and the 3' terminal is connected to a quenching group. The detection method disclosed in the invention makes of the detection kit to perform fluorescent quantitative PCR reaction.

Description

technical field [0001] The invention belongs to the fields of biotechnology and medicine, and in particular relates to a detection kit and a detection method for 19 deletion mutations of exon 19 of human EGFR gene. Background technique [0002] EGFR (Epidermal Growth Factor Receptor, Epidermal Growth Factor Receptor) gene is located in the 7p12-14 region of the short arm of human chromosome 7 and consists of 28 exons. The protein belongs to the receptor tyrosine kinase (TKI) family. EGFR is a transmembrane protein that is mainly distributed on the surface of the cell membrane. It consists of three parts: the extracellular ligand binding domain, the transmembrane region and the intracellular tyrosine kinase domain. Exons 18-24 encode the tyrosine kinase domain of the gene. amino acid kinase domain. The EGFR signaling pathway plays an important role in physiological processes such as cell growth, proliferation and differentiation. EGFR is activated by binding to ligands, inc...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68G01N21/64
Inventor 蔡从利张喆罗艳周鹏飞
Owner WUHAN YZY MEDICAL SCI & TECH
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