Single-Mismatch Position-Sensitive Detection of DNA Based on a Bifunctional Ruthenium Complex

Abstract

A ruthenium complex, pentaamine ruthenium [3-(2-phenanthren-9-yl-vinyl)-pyridine] (which we refer to as RuL in the text) generated in situ has been used as a sensitive and selective electrochemical indicator in DNA sensing. The complex incorporates dual functionalities with the Ru center providing a redox probe and the ligand (L) providing a fluorescent tag. The presence of the aromatic groups in the ligand endows the complex with an intercalative character and makes it capable of binding to double-stranded DNA (dsDNA) more efficiently than to single-stranded DNA (ssDNA). Combining spectroscopic and electrochemical techniques, we have elucidated the nature of the interactions. From these data we conclude that the binding mode is fundamentally intercalative. The ligand-based fluorescence allows characterization of the complex formation as well as for melting experiments to be carried out. The metal-based redox center is employed as an electrochemical indicator to detect the hybridization event in a DNA biosensor. The biosensor has been developed by immobilization of a thiolated capture probe sequence from Helicobacter pylori onto gold electrodes. With the use of this approach, complementary target sequences of Helicobacter can be quantified over the range of 106 to 708 pmol with a detection limit of 92+/-0.4 pmol and a linear correlation coefficient of 0.995. In addition, this approach allows the detection, without the need for a hybridization suppressor in solution, such as formamide, of not only a single mismatch but also its position in a specific sequence of H. pylori, due to the selective interaction of this bifunctional ruthenium complex with dsDNA.

Screening of Specific Gene Mutations Associated with Cystic Fibrosis

“…Sequences amplified by PCR of the DNA samples and primers used in this work. www.electroanalysis.wiley-vch.de(2-phenanthren-9-yl-vinyl)-pyridine] (which we refer as RuL), as a sensitive and selective electrochemical hybridization indicator in DNA sensing, able to single mismatch detection [29,30]. Based on the use of this ruthenium complex and an AuSPEs as a drop-on sensor we have recently developed a selective disposable amperometric DNA biosensor to detect SNP directly in large sequences obtained from PCR [30].…”

“…www.electroanalysis.wiley-vch.de(2-phenanthren-9-yl-vinyl)-pyridine] (which we refer as RuL), as a sensitive and selective electrochemical hybridization indicator in DNA sensing, able to single mismatch detection [29,30]. Based on the use of this ruthenium complex and an AuSPEs as a drop-on sensor we have recently developed a selective disposable amperometric DNA biosensor to detect SNP directly in large sequences obtained from PCR [30]. The detection relies on the comparison of the voltammetric transduction of the hybridization reaction between the immobilized probe (obtained from denaturation of a thiolated wild-type PCR sample) and target DNA sequences (wild type or mutant) present in the sample.…”

The intensification of luminol electrochemiluminescence by metallic oxide nanoparticles

“…These results suggested there was not any chemical reaction between luminol and MONPs. But the UV-vis spectra also indicated the hyperchromic or hypochromic effect [22][23][24] from the MONPs. After mixed with MONPs, the absorbances of three absorptive peaks of luminol around 222 nm, 300 nm and 348 nm are different with mathematic overlapped values (see Table 2).…”

Sol–gel derived gold nanoparticles biosensing platform for Escherichia coli detection

“…As can be seen in this table, for the biosensor developed by immobilization of the probe onto bare gold electrodes (DNA/Au), a detection limit of 92 pmol was found [35]. In a second approach [8], AuNPs were electrodeposited onto the gold electrode surface (DNA/AuNPs/Au) prior to the immobilization of the capture probe.…”

“…After exposition of this biosensing platform to a solution containing the target sequence, the hybridization event was detected using differential pulse voltammetry (DPV) from changes in the electroactivity of the redox indicator pentaamin ruthenium [3-(2-phenanthren-9-yl-vinyl)-pyridine] complex, previously incorporated into ds-DNA layer formed after hybridization. This compound is capable of binding to double-stranded DNA more efficiently than to single-stranded DNA in a mode that is fundamentally intercalative due to the presence of an aromatic group in the ligand moiety [8,[34][35][36], allowing the detection not only of a complementary Escherichia coli sequence but also the presence of a mismatched sequence.…”