A universal approach for mutation identification with a DNA probe–enzyme combination platform

Abstract

The development of rapid, and highly specific approaches for mutation detection is crucial for clinical diagnosis, since gene mutations are closely related to the progression of various diseases. In this study, we developed a convenient nucleic acid probe-promoted enzymatic reaction to assist in the sensitive detection of mutations in clinical samples. A set of DNA probes were designed to achieve preliminary mutation discrimination and subsequent qPCR steps ensured target enrichment in low abundance. The flexibility of DNA probes and specificity of the enzymatic method were investigated by detecting TP53 R273L, BRAF G469V, EGFR G719C and other types of mutations; the approach is able to accurately detect variants at allele frequencies as low as 0.01–0.1% in less than 2 hours. Around 12 mutations that are associated with the occurrence and progression of lung squamous cell carcinoma were tested. We anticipate that our strategy for the universal design principle of DNA probes towards various targets will provide a general route for sensitive, fast and cost-effective detection of rare mutations, offering great potential in clinical applications.