title: Cyclic denaturation and renaturation of double-stranded DNA by redox-state switching of DNA intercalators.
abstract: Hybridization of complementary nucleic acid strands is fundamental to nearly all molecular bioanalytical methods ranging from polymerase chain reaction and DNA biosensors to next generation sequencing. For nucleic acid amplification methods, controlled DNA denaturation and renaturation is particularly essential and achieved by cycling elevated temperatures. Although this is by far the most used technique, the management of rapid temperature changes requires bulky instrumentation and intense power supply. These factors so far precluded the development of true point-of-care tests for molecular diagnostics. To overcome this limitation we explored the possibility of using electrochemical means to control reversible DNA hybridization by using the electroactive intercalator daunomycin (DM). We show that redox-state switching of DM altered its properties from DNA binding to nonbinding, under otherwise constant conditions, and thus altered the thermodynamic stability of duplex DNA. The operational principle was demonstrated using complementary synthetic 20mer and 40mer DNA oligonucleotides. Absorbance-based melting curve analysis revealed significantly higher melting temperatures for DNA in the presence of oxidized compared to chemically reduced DM. This difference was exploited to drive cyclic electrochemically controlled denaturation and renaturation. Analysis with in situ UV-vis and circular dichroism spectroelectrochemistry, as two independent techniques, indicated that up to 80% of the DNA was reversibly hybridized. This remarkable demonstration of electrochemical control of five cycles of DNA denaturation and renaturation, under otherwise constant conditions, could have wide-ranging implications for the future development of miniaturized analytical systems for molecular diagnostics and beyond.
authors: Division of Pathway Medicine, School of Biomedical Sciences, The University of Edinburgh, Chancellor's Building, Little France Crescent, Edinburgh EH16 4SB, Scotland, UK.
authors: Shahida N
authors: SN
authors: Syed
authors: Holger
authors: H
authors: Schulze
authors: Daniel
authors: D
authors: Macdonald
authors: Jason
authors: J
authors: Crain
authors: Andrew R
authors: AR
authors: Mount
authors: Till T
authors: TT
authors: Bachmann
mesh_terms: D004247:DNA / Q000737:chemistry*; D003630:Daunorubicin / Q000737:chemistry; D015394:Molecular Structure; D021141:Nucleic Acid Amplification Techniques; D009691:Nucleic Acid Denaturation; D010084:Oxidation-Reduction; D013696:Temperature
chemical_list: D004247:DNA; D003630:Daunorubicin