HRM (high resolution melting) method and kit for clinically detecting EGFR (epidermal growth factor receptor) gene mutation

Abstract

The invention discloses an HRM (high resolution melting) method and kit for clinically detecting EGFR (epidermal growth factor receptor) gene mutation. The DNA (deoxyribonucleic acid) of a sample to be detected is extracted, and PCR (polymerase chain reaction) amplification and HRM scanning analysis are performed by use of primers shown by SEQ ID No.1-30; the type of the EGFR gene mutation is judged according to the scanning analysis result; if the HRM scanning no-parting sample is negative, the HRM scanning parting sample is positive. The kit disclosed by the invention comprises the primers, a negative control, a positive control, a PCR reagent, LCGREEN and the like. All primers of the kit can perform PCR reaction at the same temperature, the detection sensitivity is high, the specificity is strong, the reaction system is small, the detection sample is widely available, the reaction is simple and quick, the reagent is cheap, and the flux is high. By adopting the method and kit disclosed by the invention, the whole detection process is simple to operate, time-saving, low in cost, suitable for quickly detecting the EGFR gene mutation of clinical tumor patients, and good for guiding the clinical individualized medication.

Description

technical field [0001] The invention relates to the technical field of biological detection, in particular to an HRM method and a kit for clinical detection of EGFR gene mutation. Background technique [0002] Epidermal Growth Factor (EFG) is one of the important members of the growth factor family. It binds to Epidermal Growth Factor Receptor (EGFR) and initiates downstream signaling pathways to play a series of physiological and pathological functions. effect. EGFR is a transmembrane glycoprotein composed of 1186 amino acids and a molecular weight of 170kD. Its structure includes an extracellular binding region, a transmembrane region, and an intracellular region mainly composed of tyrosine kinases. EGFR is mainly distributed on the cell membrane surface, belongs to the type I tyrosine kinase receptor family, and has tyrosine kinase activity. EGFR activation activates multiple downstream signal transduction pathways, such as RAS-RAF-MEK-MAPK pathway, PI3K-Akt pathway, PK...

AI Technical Summary

Problems solved by technology

At present, the patent invented by Suzhou Weizhen Biotechnology Co., Ltd. involves the detection of EGFR gene mutation by HRM method, but its detection accuracy is relatively low (3 out of 40 pairs of samples were found to have inconsistent results), and the consumption of reagents is large (20μL reaction system). The reaction time is long, and the results are not intuitive (critically positive is difficult to distinguish), etc.; the fluorescent quantitative PCR method has the disadvantages of high sample requirements, high reagent costs, and long reaction time.

With the increasing number of clinical specimens, the existing clinical EGFR gene mutation detection methods can no longer meet the needs, and there is an urgent need for a fast, accurate, easy-to-operate, high-sensitivity EGFR gene detection technology that avoids cross-contamination

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