A Novel Quantitative Electrochemical Method to Monitor DNA Double-Strand Breaks Caused by a DNA Cleavage Agent at a DNA Sensor
| Autor: | John Colleran, Anna Banasiak, John Cassidy |
|---|---|
| Přispěvatelé: | Technological University Dublin, Fiosraigh Award |
| Rok vydání: | 2018 |
| Předmět: |
Base pair
Biomedical Engineering Biophysics Antineoplastic Agents 010402 general chemistry Cleavage (embryo) 01 natural sciences DNA cleavage agent chemistry.chemical_compound Capillary electrophoresis Cleave Electrochemistry DNA quantitation A-DNA DNA Breaks Double-Stranded Nuclease DNA cleavage biology Oligonucleotide Mutagenicity Tests 010401 analytical chemistry General Medicine Electrochemical Techniques Combinatorial chemistry Copper bis-phenanthroline 0104 chemical sciences DNA interaction Chemistry chemistry biology.protein DNA Copper DNA biosensor Biotechnology |
| Zdroj: | Articles |
| Popis: | To date, DNA cleavage, caused by cleavage agents, has been monitored mainly by gel and capillary electrophoresis. However, these techniques are time-consuming, non-quantitative and require gel stains. In this work, a novel, simple and, importantly, a quantitative method for monitoring the DNA nuclease activity of potential anti-cancer drugs, at a DNA electrochemical sensor, is presented. The DNA sensors were prepared using thiol-modified oligonucleotides that self-assembled to create a DNA monolayer at gold electrode surfaces. The quantification of DNA double-strand breaks is based on calculating the DNA surface coverage, before and after exposure to a DNA cleavage agent. The nuclease properties of a model DNA cleavage agent, copper bis-phenanthroline ([CuII(phen)2]2+), that can cleave DNA in a Fenton-type reaction, were quantified electrochemically. The DNA surface coverage decreased on average by 21% after subjecting the DNA sensor to a nuclease assay containing [CuII(phen)2]2+, a reductant and an oxidant. This percentage indicates that 6 base pairs were cleaved in the nuclease assay from the immobilised 30 base pair strands. The DNA cleavage can be also induced electrochemically in the absence of a chemical reductant. [CuII(phen)2]2+ intercalates between DNA base pairs and, on application of a suitable potential, can be reduced to [CuI(phen)2]+, with dissolved oxygen acting as the required oxidant. This reduction process is facilitated through DNA strands via long-range electron transfer, resulting in DNA cleavage of 23%. The control measurements for both chemically and electrochemically induced cleavage revealed that DNA strand breaks did not occur under experimental conditions in the absence of [CuII(phen)2]2+. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|