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In Vitro Isolation of Small-Molecule-Binding Aptamers With Intrinsic Dye-Displacement Functionality

NCJ Number
Nucleic Acids Research Volume: 46 Issue: 8 Dated: 2018
Date Published
7 pages
This article reports on a project that developed a simple strategy for directly isolating signal-reporting aptamers in vitro through systematic evolution of ligands by exponential enrichment (SELEX) that transduce binding events into a detectable change of absorbance via target-induced displacement of a small-molecule dye.

Aptamer-based sensors offer a powerful tool for molecular detection, but the practical implementation of these biosensors is hindered by costly and laborious sequence engineering and chemical modification procedures. The current project first demonstrated that diethylthiatricarbocyanine (Cy7) can stack into DNA three-way junctions (TWJs) in a sequence-independent fashion, greatly altering the dye's absorbance spectrum. The project then designed a TWJ-containing structured library and isolated an aptamer against 3,4-methylenedioxypyrovalerone (MDPV), a synthetic cathinone that is an emerging drug of abuse. This aptamer intrinsically binds Cy7 within its TWJ domain, but MDPV efficiently displaces the dye, resulting in a change in absorbance within seconds. This assay is label-free and detects nanomolar concentrations of MDPV. It also recognizes other synthetic cathinones, offering the potential to detect newly-emerging designer drugs, but does not detect structurally-similar non-cathinone compounds or common cutting agents. Moreover, the project demonstrated that the Cy7-displacement colorimetric assay is more sensitive than a conventional strand-displacement fluorescence assay. The researchers believe this strategy offers an effective generalized approach for the development of sensitive dye-displacement colorimetric assays for other small-molecule targets. (publisher abstract modified)

Date Published: January 1, 2018