Amplified detection of single base mismatches with the competing-strand assay reveals complex kinetic and thermodynamic behavior of strand displacement at the electrode surface

Katarzyna Biała; Ada Sedova; Maren Mix; Katharina Bär; Petr Orsag; Miroslav Fojta; Gerd Uwe Flechsig

doi:10.1016/j.electacta.2018.07.188

Amplified detection of single base mismatches with the competing-strand assay reveals complex kinetic and thermodynamic behavior of strand displacement at the electrode surface

Katarzyna Biała, Ada Sedova, Maren Mix, Katharina Bär, Petr Orsag, Miroslav Fojta, Gerd Uwe Flechsig

Research output: Contribution to journal › Article › peer-review

5 Scopus citations

Abstract

Detection of single-base mismatches with respect to a probe strand has been a predominant pursuit in electrochemical biosensor efforts, due to links found between single nucleotide polymorphisms (SNPs) and the predisposition to various diseases. We report an osmium tetroxide bipyridine-based, thermally-controlled, competitive-strand electrochemical assay to allow amplified detection of single-base mismatches. Optimally designed competitive-strand displacement and hybridization temperature allows us to distinguish the single-mismatched-target from the fully complementary sequence with unambiguous, highly reproducible, robust signal differences of over 90%. Furthermore, we find a complex interplay between the position of the redox label, variations in strand displacement kinetics due to mismatches incorporated into the competitive strand, and alterations in the melting temperature of DNA duplexes tethered on the gold surface, when probed by square-wave voltammetry. These insights will apply to any surface-tethered DNA-based electrochemical biosensor, and can help with understanding complex phenomena involved in these types of assays.

Original language	English
Pages (from-to)	272-283
Number of pages	12
Journal	Electrochimica Acta
Volume	285
DOIs	https://doi.org/10.1016/j.electacta.2018.07.188
State	Published - Sep 20 2018
Externally published	Yes

Funding

The authors are grateful to SUNY-Albany for research start-up funding, as well as Prof. Dr. med. Ibrahim Saleh and Dr. Lemcke of the University at Lübeck and Immungenetics AG, Lübeck, Germany, for advice regarding SNPs with relevance to autoimmune diseases. The authors also thank the German Research Foundation (DFG Heisenberg Fellowship FL 384/7-2), as well as the German Federal Ministry for Education and Research (BMBF WK Potenzial grant 03WKP17A “Nordic Diagnostic Alliance”) for financial support. M.F and P.O. were supported by the Czech Science Foundation ( P206/12/G151 ) and by the ASCR ( RVO 68081707 ). This research used resources of the Oak Ridge Leadership Computing Facility, which is a DOE Office of Science User Facility supported under Contract DE-AC05-00OR227525 .

Funders	Funder number
California Department of Fish and Game	FL 384/7-2
Office of Science	DE-AC05-00OR227525
Advanced Scientific Computing Research	RVO 68081707
Deutsche Forschungsgemeinschaft
Grantová Agentura České Republiky	P206/12/G151
Bundesministerium für Bildung und Forschung	03WKP17A

Keywords

Melting temperature
Osmium tetroxide bipyridine labeling
Single-base mismatch detection
Surface-immobilized DNA

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10.1016/j.electacta.2018.07.188

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title = "Amplified detection of single base mismatches with the competing-strand assay reveals complex kinetic and thermodynamic behavior of strand displacement at the electrode surface",

abstract = "Detection of single-base mismatches with respect to a probe strand has been a predominant pursuit in electrochemical biosensor efforts, due to links found between single nucleotide polymorphisms (SNPs) and the predisposition to various diseases. We report an osmium tetroxide bipyridine-based, thermally-controlled, competitive-strand electrochemical assay to allow amplified detection of single-base mismatches. Optimally designed competitive-strand displacement and hybridization temperature allows us to distinguish the single-mismatched-target from the fully complementary sequence with unambiguous, highly reproducible, robust signal differences of over 90%. Furthermore, we find a complex interplay between the position of the redox label, variations in strand displacement kinetics due to mismatches incorporated into the competitive strand, and alterations in the melting temperature of DNA duplexes tethered on the gold surface, when probed by square-wave voltammetry. These insights will apply to any surface-tethered DNA-based electrochemical biosensor, and can help with understanding complex phenomena involved in these types of assays.",

keywords = "Melting temperature, Osmium tetroxide bipyridine labeling, Single-base mismatch detection, Surface-immobilized DNA",

author = "Katarzyna Bia{\l}a and Ada Sedova and Maren Mix and Katharina B{\"a}r and Petr Orsag and Miroslav Fojta and Flechsig, {Gerd Uwe}",

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AU - Biała, Katarzyna

AU - Sedova, Ada

AU - Mix, Maren

AU - Bär, Katharina

AU - Orsag, Petr

AU - Fojta, Miroslav

AU - Flechsig, Gerd Uwe

PY - 2018/9/20

Y1 - 2018/9/20

N2 - Detection of single-base mismatches with respect to a probe strand has been a predominant pursuit in electrochemical biosensor efforts, due to links found between single nucleotide polymorphisms (SNPs) and the predisposition to various diseases. We report an osmium tetroxide bipyridine-based, thermally-controlled, competitive-strand electrochemical assay to allow amplified detection of single-base mismatches. Optimally designed competitive-strand displacement and hybridization temperature allows us to distinguish the single-mismatched-target from the fully complementary sequence with unambiguous, highly reproducible, robust signal differences of over 90%. Furthermore, we find a complex interplay between the position of the redox label, variations in strand displacement kinetics due to mismatches incorporated into the competitive strand, and alterations in the melting temperature of DNA duplexes tethered on the gold surface, when probed by square-wave voltammetry. These insights will apply to any surface-tethered DNA-based electrochemical biosensor, and can help with understanding complex phenomena involved in these types of assays.

AB - Detection of single-base mismatches with respect to a probe strand has been a predominant pursuit in electrochemical biosensor efforts, due to links found between single nucleotide polymorphisms (SNPs) and the predisposition to various diseases. We report an osmium tetroxide bipyridine-based, thermally-controlled, competitive-strand electrochemical assay to allow amplified detection of single-base mismatches. Optimally designed competitive-strand displacement and hybridization temperature allows us to distinguish the single-mismatched-target from the fully complementary sequence with unambiguous, highly reproducible, robust signal differences of over 90%. Furthermore, we find a complex interplay between the position of the redox label, variations in strand displacement kinetics due to mismatches incorporated into the competitive strand, and alterations in the melting temperature of DNA duplexes tethered on the gold surface, when probed by square-wave voltammetry. These insights will apply to any surface-tethered DNA-based electrochemical biosensor, and can help with understanding complex phenomena involved in these types of assays.

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Amplified detection of single base mismatches with the competing-strand assay reveals complex kinetic and thermodynamic behavior of strand displacement at the electrode surface

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Keywords

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