Quantitative Measurement of Electrochemically Induced DNA Damage Using Capillary Electrophoresis

Published

May 3, 2013

Author(s)

Donald H. Atha, Niyatie Ammanamanchi, Mofiyin Obadina, Vytautas Reipa

Abstract

Exposure of mammalian cells to oxidative stress can result in DNA damage that adversely affects many cell processes. We used bulk electrolysis in an electrochemical system and capillary electrophoresis (CE) to control and measure the effect of oxidative stress on DNA. Calf thymus DNA and fluorescently labeled DNA sizing ladder were subjected to fixed oxidizing potentials using a reticulated vitreous carbon electrode (RVC) and their fragmentation was measured with the use of CE. The resulting electropherograms showed that the oxidative treatment only resulted in DNA fragmentation. Poly adenosine (Poly A) 40mer and poly guanosine (Poly G) 40mer oligonucleotides were exposed to a controlled oxidative environment at constant potential values E = 0.5V, 1.0V, 1.5V and 2V (vs Ag/AgCl) for 1 hour in 0.1 mol/L K phosphate buffer pH 7.3. The treated DNA fragments were analyzed by CE. The areas of the CE peaks were measured and the percentage of DNA fragmentation was calculated. Only minor fragmentation was observed while oligonucleotides were exposed to E = 0.5V, with strand scission starting at electrode potentials E > 1.0V. The E = 2.0V treatment resulted in approximately 50% fragmentation of Poly A, compared to approximately 15% for the Poly G. These results, using DNA as a test model, demonstrate that controlled-potential electrolysis can be used to produce desired levels of oxidative stress to biomaterial without added chemicals, which would serve as quantitative test vehicle for nucleic acid stability thermodynamic models.

Citation

Journal of the Electrochemical Society

Pub Type

Journals

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DOI Link

Keywords

Electrochemical oxidation, Reticulated carbon electrode, Oxidative DNA damage, DNA strand scission, Capillary electrophoresis

Citation

Atha, D. , Ammanamanchi, N. , Obadina, M. and Reipa, V. (2013), Quantitative Measurement of Electrochemically Induced DNA Damage Using Capillary Electrophoresis, Journal of the Electrochemical Society, [online], https://doi.org/10.1149/2.021307jes (Accessed December 4, 2024)