A cost-effective detection of low-abundance mutation with DNA three-way junction structure and lambda exonuclease

We presented a low-abundance mutation detection method with lambda exonuclease and DNA three-way junction structure. The assistant strand in the DNA three-way junction structure could regulate the reaction system from the kinetics and thermodynamics aspects. The optimization of the assistant strand helps to improve the selectivity of the mutant-type DNA to the wild-type DNA about 35 times. Moreover, the cost of the optimization process could be saved by about 90%. The method was applied to the detection of a human ovarian cancer-related gene mutation BRCA1 (rs1799949, c.2082C>T). The limit of detection to the mutation abundance in the DNA three-way junction structure system (0.2%) was one order lower compared with that in the double-stranded DNA structure system (2%). The mutation abundance in different standard samples was quantitively measured, and the results were consistent with the initial abundance in the standard samples. (C) 2020 Chinese Chemical Society and Institute of Materia Medica, Chinese Academy of Medical Sciences. Published by Elsevier B.V. All rights reserved.