G-quadruplex specific dye-based ratiometric FRET aptasensor for robust and ultrafast detection of toxin

Abstract

G-quadruplex specific dyes are powerful tools for probing nucleic acid structures. Among nucleic acids, aptamers are of great interest, and widely exploited to construct versatile bioassays. Herein, based on G-quadruplex selective dye, thioflavin T (ThT), for probing the intrinsic structure of aptamers, we proposed a ratiometric fluorescence resonance energy transfer (FRET) aptasensor enabling robust and ultrafast detection of toxin. The binding of target ochratoxin A (OTA) would destruct the G-quadruplex structure of aptamer. It would lead to the detachment of ThT dye from aptamer which diminished the FRET effect between ThT and terminal-labeled dye, thus allowing quantification of OTA via FRET signals. The FRET aptasensor would confer an enhancement of 76.9% of signal to background ratio compared to the ThT-based non-FRET aptasensor. Remarkably, the FRET mechanism would eliminate the signal fluctuation resulted from varied probe concentration, thus benefiting the robustness of the assay. The aptasensor could achieve a detection of limit of 0.38 ng/mL for OTA detection. And the detection of OTA could be finished within 30 s. Besides, the assay was successful in analyzing OTA in coffee and oat samples with recoveries rate of 93.93%–107.59%. Therefore, G-quadruplex specific dye-based probing and FRET method would be a compelling design strategy for aptasensor, and may facilitate their practical application in food safety and environmental screening.