Target-assembled tandem oligonucleotide systems based on exciplexes for detecting DNA mismatches and single nucleotide polymorphisms

Abstract

We report the first exciplex-based split-probe system for DNA detection. The detector is split at a molecular level into signal-silent components which, before a signal is generated, must be assembled correctly into a particular three-dimensional arrangement. The model system comprises of two 8-mer oligonucleotides, complementary to neighbouring sites of a 16-mer DNA target, each equipped with moieties able to form an exciplex on correct, contiguous hybridization. The exciplex emits at approximately 480 nm with a large Stokes shift (135 nm). The extremely rigorous structural demands for exciplex formation and emission were achieved by careful structural design and by the discovery that high levels of certain organic solvents (especially trifluoroethanol) specifically favour emission of the DNA-mounted exciplex, probably the net result of the particular duplex structure and specific solvation of the exciplex partners. Inserts and mismatches can be effectively detected by this exciplex construct giving potential for single nucleotide polymorphism detection.