Original article
English
Gbaj A1, 2, Hawisa1 S, Walsh L2, Rogert C 2, 3, Sardarian A4, Bichenkova EV2, Etchells LL2, Douglas KT2
1-National Centre for Medical Research (El-Zawia, Libya). 2-Wolfson Centre for Rational Structure-Based Design of Molecular Diagnostics, School of Pharmacy and Pharmaceutical Sciences, University of Manchester, Manchester, M13 9PL, U.K. 3- Chesterford Research Park, Little Chesterford, Essex, CB10 1XL, U. K. 4-Department of Chemistry, University of Shiraz, Shiraz, Iran
JMJ 2009;9(4):250-257
Abstract
The development of a new sensitive, safe and easy to use exciplex probes system for the detection of DNA is of relevant importance. The aim of this study was to develop a fluorescence- based technique to work in homogenous, and then heterogeneous assays systems. Exciplex signal with DNA hybridisation were determined in solution using fluorescence measurements and complementary fluorophore-labeled oligonucleotides. One oligonucleotide probe, was labelled with a 5′-terminal pyrene, and the other was labelled with 3′-terminal naphthalene. The juxtaposition of the two labels in double-stranded complexes, results in a strong exciplex signal at a longer wavelength (~ 480nm) thereby providing the means to differentiate single-stranded DNA from double-stranded DNA. Since measurements were based on fluorescence, DNA denaturation and association could be monitored routinely at low DNA concentrations. The importance of the presence of various counter cations (Na+, K+ and Mg2+) on exciplex signal intensity was evaluated. The results obtained represent the first case of an oligonucleotide terminally located probe system based on exciplex fluorescence emission.
Keywords: Terminally located probe systems, Exciplex; DNA detection, Fluorescence, Stokes shift.
Link/DOI: http://www.jmj.org.ly/PDF/winter2009/250.pdf