Cancer gene mutation and gene amplification detection

Abstract

The invention provides a primer set for simultaneously detecting mutation of a plurality of regions of two or more exons of one or more cancer drive genes in a sample on the basis of multiplex PCR (polymerase chain reaction) and high-flux sequencing techniques, a kit comprising the primer set, and a method for amplifying the cancer drive gene or detecting the cancer drive gene by using the primer set and / or kit. The mutation detection result can be used for instructing cancer clinic medication, auxiliary diagnosis or prognosis judgment on certain cancers.

Description

field of invention [0001] The invention relates to the detection of genes with high incidence of cancer mutations. The invention provides a primer set, a kit and a detection method for multigene and multisite mutation detection of cancer samples based on multiplex PCR and high-throughput sequencing technology. The provided test results can be used, for example, to assist diagnosis or guide clinical medication regimens. Background technique [0002] Cancer is one of the most important non-communicable diseases in the world, and it is also the chronic disease with the highest fatality rate. In my country, the number of deaths due to cancer is as high as 2.7 million every year, among which lung cancer, gastric cancer, colorectal cancer and other digestive tract tumors are the most serious. Gynecological tumors, led by breast cancer, are also the leading cause of cancer death in women. According to statistics, the incidence rate of lung cancer in my country is 53.57 / 100,000, an...

AI Technical Summary

Problems solved by technology

The fluorescent PCR method has the characteristics of high sensitivity, and the technology is mature and widely used. However, each pair of primers can only detect one mutation, and each mutation requires a separate PCR reaction system, resulting in a large amount of samples and cannot be detected at the same time. A large number of samples or sites; the first-generation Sanger sequencing method can detect multiple mutations with a single pair of primers, and has the characteristics of low cost, but only one pair of primers can be used for each amplification, which cannot detect multiple genes or multiple samples at the same time, and Large sample volume required and unsatisfactory detection of infrequent mutations

[0013] The traditional classic method for detecting gene amplification is FISH (fluorescence in situ hybridization), which has good sensitivity, and its biggest advantage is that it can visually observe the gene amplification at the nucleus level, and can be detected by the brightness of fluorescence. The strength of the gene is used to estimate the number of gene copies, but this method is costly and has high requirements for the operation of technicians, and its result judgment is easily affected by many factors, such as polyploid cells, etc.

[0014] Multiplex PCR technology is currently widely used in the amplification of multiple genes and multiple sites. Compared with ordinary PCR, it also has the advantages of high efficiency, quantitative or qualitative templates, etc., but its system construction needs to integrate the homology of the target sequence. Factors such as sex and length, primer kinetics, optimization of PCR reaction conditions, etc. For example, the length range of the target sequence is too large, which may lead to uneven amplification efficiency, improper primer concentration ratio, or the formation of dimers between primers can affect The overall amplification efficiency or amplification uniformity is generally less than five to six pairs of primers in the same system on the market or in experiments at the same time, which shows that it is quite difficult and demanding in terms of technical realization

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