title: Electrochemical strategy for sensing DNA methylation and DNA methyltransferase activity.
abstract: The present work demonstrates a novel signal-off electrochemical method for the determination of DNA methylation and the assay of methyltransferase activity using the electroactive complex [Ru(NH3)6](3+) (RuHex) as a signal transducer. The assay exploits the electrostatic interactions between RuHex and DNA strands. Thiolated single strand DNA1 was firstly self-assembled on a gold electrode via Au-S bonding, followed by hybridization with single strand DNA2 to form double strand DNA containing specific recognition sequence of DNA adenine methylation MTase and methylation-responsive restriction endonuclease Dpn I. The double strand DNA may adsorb lots of electrochemical species ([Ru(NH3)6](3+)) via the electrostatic interaction, thus resulting in a high electrochemical signal. In the presence of DNA adenine methylation methyltransferase and S-adenosyl-l-methionine, the formed double strand DNA was methylated by DNA adenine methylation methyltransferase, then the double strand DNA can be cleaved by methylation-responsive restriction endonuclease Dpn I, leading to the dissociation of a large amount of signaling probes from the electrode. As a result, the adsorption amount of RuHex reduced, resulting in a decrease in electrochemical signal. Thus, a sensitive electrochemical method for detection of DNA methylation is proposed. The proposed method yielded a linear response to concentration of Dam MTase ranging from 0.25 to 10UmL(-1) with a detection limit of 0.18UmL(-1) (S/N=3), which might promise this method as a good candidate for monitoring DNA methylation in the future.
authors.affiliation: Key Laboratory of Eco-environments in Three Gorges Reservoir Region (Ministry of Education), School of Chemistry and Chemical Engineering, Southwest University, Chongqing 400715, China.
authors.forename: Gang Lin
authors.initials: GL
authors.lastname: Wang
authors.forename: Long Yin
authors.initials: LY
authors.lastname: Zhou
authors.forename: Hong Qun
authors.initials: HQ
authors.lastname: Luo
authors.forename: Nian Bing
authors.initials: NB
authors.lastname: Li
mesh_terms: D056831:Coordination Complexes / Q000737:chemistry; D004247:DNA / Q000032:analysis* / Q000737:chemistry; D019175:DNA Methylation; D055664:Electrochemical Techniques; D004566:Electrodes; D009693:Nucleic Acid Hybridization; D012428:Ruthenium / Q000737:chemistry; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific) / Q000378:metabolism; D055672:Static Electricity
chemical_list: D056831:Coordination Complexes; D012428:Ruthenium; D004247:DNA; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific)