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Biosens Bioelectron. 2018 Oct 15;117:444-449. doi: 10.1016/j.bios.2018.06.040. Epub 2018 Jun 22.

Picomolar sensitive and SNP-selective "Off-On" hairpin genosensor based on structure-tunable redox indicator signals.

Author information

1
Interdisciplinary Nanoscience Center (iNANO), Science and Technology, Aarhus University, Gustav Wieds Vej 1590-14, DK-8000 Aarhus C, Denmark.
2
Department of Molecular Medicine, Aarhus University Hospital, Palle Juul-Jensens Boulevard 99, DK-8200 Aarhus N, Denmark.
3
Interdisciplinary Nanoscience Center (iNANO), Science and Technology, Aarhus University, Gustav Wieds Vej 1590-14, DK-8000 Aarhus C, Denmark. Electronic address: elena.ferapontova@inano.au.dk.

Abstract

A robust and sensitive electrochemical assay for chrononocoulometric detection of nucleic acids at a single nucleotide polymorphism (SNP) level has been developed. The assay exploits hybridization-induced conformational switching of gold-tethered TP53-specific 33-mer and truncated 20-mer hairpin DNA probes and methylene blue (MB) as an intercalating redox indicator. We show that by fine tuning of MB-DNA intercations the enhanced binding of MB to hybrids formed with a cancer-biomarker sequence can be achieved, and that results in robust "off-on" sensing of hybridization, while the stem-loop probe design allows minimized, independent of the DNA length background signals. Both DNA probes were sensitive to the presence of SNP in the targeted DNA sequence already at 10 pM. DNA levels, and the robust "off-on" discrimination of 10 pM perfectly-matched DNA from 50 nM SNP-containing DNA was achieved by time-adjusted chronocoulometry. This label-free hairpin DNA strategy allows systematic design of DNA assays for fast, robust and inexpensive genetic analysis in excessive mixtures of structurally-related DNA sequences and was used for specific analysis of prostate-cancer-realted cellular microRNA in total RNA samples isolated from LNCaP and BPH1 cells.

KEYWORDS:

Chronocoulometry; DNA hairpin probes; Genosensor; Hybridization; Methylene Blue; Single Nucleotide Polymorphism

PMID:
29975881
DOI:
10.1016/j.bios.2018.06.040
[Indexed for MEDLINE]

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