Abstract
There is significant interest in developing methods that visualize and detect RNA. Bioorthogonal template-driven tetrazine ligations could be a powerful route to visualizing nucleic acids in native cells, yet past work has been limited with respect to the diversity of fluorogens that can be activated via a tetrazine reaction. Herein we report a novel bioorthogonal tetrazine uncaging reaction that harnesses tetrazine reactivity to unmask vinyl ether caged fluorophores spanning the visible spectrum, including a near-infrared (NIR)-emitting cyanine dye. Vinyl ether caged fluorophores and tetrazine partners are conjugated to high-affinity antisense nucleic acid probes, which show highly selective fluorogenic reactivity when annealed to their respective target RNA sequences. A target sequence in the 3' untranslated region of an expressed mRNA was detected in live cells employing appropriate nucleic acid probes bearing a tetrazine-reactive NIR fluorogen. Given the expansion of tetrazine fluorogenic chemistry to NIR dyes, we believe highly selective proximity-induced fluorogenic tetrazine reactions could find broad uses in illuminating endogenous biomolecules in cells and tissues.
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