Detection of miRNA in live cells by using templated RuII- catalyzed unmasking of a fluorophore

Abstract

Reactions templated by cellular nucleic acids are attractive for nucleic acid sensing or responsive systems. Herein we report the use of a photocatalyzed reductive cleavage of an immolative linker to unmask a rhodamine fluorophore, and its application to miRNA imaging. The reaction was found to proceed with a very high turnover (>4000) and provided reliable detection down to 5 pM of template by using γ-serine-modified peptide nucleic acid (PNA) probes. The reaction was used for the selective detection of miR-21 in BT474 cells and miR-31 in HeLa cells following irradiation for 30 min. The probes were introduced by using reversible permeation with streptolysin-O (SLO) or a transfection technique. Seeing is believing: Different miRNAs were used as templates for the photoreduction of an azide-based immolative linker by Ru II-peptide nucleic acid (PNA) conjugates to uncage rhodamine (see figure). The method was validated by using two sets of γ-serine-modified PNA derivatives with varying affinities to their target (perfect match and mismatch). Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.