Kit and detection method for detection of human KRAS gene mutations

Abstract

The invention discloses a kit for detection of human KRAS gene mutations. The kit comprises specific primer pairs of seven genes and seven probe primers cooperatively used with the specific primers. Mixed liquid of primers and probes includes mutation site genes and internal-control primer probes, quality control genes and internal-control detection primer probes, FAM fluorescein used for labeling mutation and quality control probes and an internal control marker HEX fluorescein; quality control and internal control are taken into consideration in result interpretation; the quality control genes are selected from the conserved segment of the human KRAS gene, and the conserved human beta-actin gene is used as internal control; and a double-control system is employed for monitoring the mass of DNA and PCR reaction process in blood plasma samples. Compared with the prior art, the invention has the following beneficial effects: 1) the method has high sensitivity and can detect 1% trace mutation templates under the background of 100 ng of wild mankind genomes, so false negative results are reduced; 2) a totally-enclosed reaction is carried out to avoid false positive results; and 3) the double-control reaction system is employed, so PCR amplification and nucleic acid extraction are effectively detected via quality control and internal control.

Description

technical field [0001] The invention relates to the fields of biotechnology and medicine, in particular to a kit for detecting human KRAS gene mutation and a detection method thereof. Background technique [0002] At present, lung cancer has become the leading cause of cancer death in the world, with 1.5 million new cases each year worldwide, seriously threatening human health. Among lung cancer patients, non-small cell lung cancer (NSCLC) accounts for about 80%, and 80% of them have lost the best chance of surgery or radiotherapy at the time of treatment, resulting in low long-term survival rate and worrying efficacy. The 2-year survival rate of stage IIIB / IV non-small cell lung cancer is only 10%-20%, and the median survival time is only 10-12 months. Therefore, people need to continuously study new treatment methods to improve the effectiveness of lung cancer treatment. Among them, new treatment methods represented by molecular targeted therapy have brought new hope for ...

AI Technical Summary

Problems solved by technology

Therefore, the extracted DNA often contains a large amount of wild-type DNA, so the detection of somatic mutations requires high specificity, and the current widely used direct sequencing method has limited detection capabilities and cannot fully meet clinical needs.

Specific probe methods such as the UK DxS represented by Scorpions ARMS and the KRAS detection kit of Xiamen Adx in China have high detection sensitivity, but their detection costs are high, and they are not suitable for routine detection and screening of domestic NSCLC patients. Therefore, clinical There is an urgent need to develop a fast, accurate, easy-to-operate, and highly sensitive kit for detecting KRAS gene mutations to meet the timeliness requirements of clinical medication and detection and diagnosis

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