Fluorescent DNA base modifications and substitutes: Multiple fluorophore labeling and the DETEQ concept

There is an increasing need for fluorescent nucleic acid probes that are able to sense genetic variations without the application of enzymes. The incorporation of organic fluorophores either as DNA base modifications or as DNA base substitutions represents a powerful and versatile method for such new fluorescent DNA assays. Multiple labeling of oligonucleotides using several adjacent chromophore-modified DNA bases yields fluorescence enhancement and modulation that are sensitive to single-base mismatches in the complementary oligonucleotide. Charge transfer processes that cause fluorescence quenching are DNA-base mediated and occur over several base pairs distance. Our DETEQ setup, consisting of a fluorescence DNA base substitution and the charge acceptor as a second modification two base pairs away, allows the homogeneous detection of single-base mutations simply by fluorescence readout. This could lead to new DNA microarrays which are based on charge transfer processes and can be analyzed by commonly used fluorescence readout techniques.