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Proceedings Paper

Self-quenching DNA probes based on aggregation of fluorescent dyes
Author(s): Gabriela Schafer; Matthias Muller; Bernhard Hafner; Gregor Habl; Oliver Nolte; Nicole Marme; Jens-Peter Knemeyer
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Paper Abstract

Here we present a novel class of self-quenching, double-labeled DNA probes based on the formation of non fluorescent H-type dye dimers. We therefore investigated the aggregation behavior of the red-absorbing oxazine derivative MR121 and found a dimerization constant of about 3000 M-1. This dye was successfully used to develop hairpin-structured as well as linear self-quenching DNA probes that report the presence of the target DNA by an increase of the fluorescence intensity by a factor of 3 to 12. Generally fluorescence quenching of the hairpin-structure probes is more efficient compared to the linear probes, whereas the kinetic of the fluorescence increase is significantly slower. The new probes were used for the identification of different mycobacteria and their antibiotic resistant species. As a test system a probe for the identification of a DNA sequence specific for the Mycobacterium xenopi was synthesized differing from the sequence of the Mycobacterium fortuitum by 6 nucleotides. Furthermore we developed a method for the discrimination between the sequences of the wild type and an antibiotic resistant species of Mycobacterium tuberculosis. Both sequences differ by just 2 nucleotides and were detected specifically by the use of competing olignonucleotides.

Paper Details

Date Published: 4 April 2005
PDF: 10 pages
Proc. SPIE 5704, Genetically Engineered and Optical Probes for Biomedical Applications III, (4 April 2005); doi: 10.1117/12.588257
Show Author Affiliations
Gabriela Schafer, Univ. Heidelberg (Germany)
Matthias Muller, Hygieneinstitute (Germany)
Bernhard Hafner, Hygieneinstitute (Germany)
Gregor Habl, Deutsches Krebsforschungszentrum (Germany)
Oliver Nolte, Hygieneinstitute (Germany)
Nicole Marme, Univ. Heidelberg (Germany)
Jens-Peter Knemeyer, Deutsches Krebsforschungszentrum (Germany)

Published in SPIE Proceedings Vol. 5704:
Genetically Engineered and Optical Probes for Biomedical Applications III
Darryl J. Bornhop; Samuel I. Achilefu; Ramesh Raghavachari; Alexander P. Savitsky, Editor(s)

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