An exonuclease III-amplified 4-way strand migration system for low-abundance deletion mutation

A 4-way strand exchange competitive DNA testing system based on Holliday junction has an advantage in realizing high sensitivity and specificity simultaneously. However, the kinetics is limited without enzyme assisting. Herein, we constructed a method that combined a 4-way strand migration system and exonuclease III (Exo III). For the properties of Exo III that has high catalytic effects and no specific recognition site, a DNA probe assisted by Exo III is easy to design and synthesize. We applied the system to detect different lengths of deletion mutation, and the results showed that the time to differentiate wild-type DNA and mutant-type DNA was so short within 5-20 min. Besides, the discrimination factor (DF) was as high as 1177.88 for EGFR-15-nt deletion, and the mutation detection limit was as low as 0.02% for PBRM1-8-nt deletion. Without adding any other specific label, the Exo III-amplified 4-way strand migration system is a simple, sensitive, selective, and cost-effective method that suggests a potential possibility for the diagnosis of cancers.