Multiplexed DNA detection with DNA tweezers in a one-pot reaction

Abstract

One of the great challenges for the development of biosensors is to construct a fast, practical and multiplexed detection platform. One promising approach is utilizing the controllable conformation of DNA nanostructures for such biosensing applications. Here, we present tweezer-like DNA (TW-DNA)-assisted multiplexed DNA detection in a one-pot reaction based on fluorescence quenching. The open conformation of the TW-DNA structure can be changed to a closed state by a specific single-stranded DNA target. The fluorophore/quencher pairs decorated on the TW-DNA and the conformation changes were observed by fluorescence quenching. In its initial state, the TW-DNA is maintained in an open state that demonstrates high fluorescence intensity. In the presence of a specific target DNA, the TW-DNA changes to a closed conformation and the emission intensity of the donor is quenched which is directly proportional to the target concentration. Three DNA targets were simultaneously detected using three individually self-assembled TW-DNA that were modified appropriately with fluorophore/quencher pairs for the specific detection of the individual DNA target. The detection range of this assay is at submicromolar concentration of multiple DNA targets with a fast detection time of one minute. Moreover, this biosensor platform provides the significant advantage of being a one-pot reaction for multiplexed DNA detection without any cross-talk, thereby making it a highly applicable assay. Keywords: Multiplexing, DNA detection, Fluorescence quenching, One-pot detection, DNA nanostructure