Issue 8, 2009
From the journal:

Analyst

Hybridization kinetics of double-stranded DNA probes for rapid molecular analysis†

Vinay Gidwani,a   Reza Riahi,b   Donna D. Zhangc  and  Pak Kin Wong*bd  

* Corresponding authors

a Department of Biochemistry and Molecular Biophysics, University of Arizona, Tucson, Arizona, USA

b Department of Aerospace and Mechanical Engineering, University of Arizona, Tucson, Arizona, USA
E-mail: pak@email.arizona.edu
Fax: +1-520-621-8191
Tel: +1-520-626-2215

c Department of Pharmacology and Toxicology, University of Arizona, Tucson, Arizona, USA

d Biomedical Engineering Interdisciplinary Program and Bio5 Institute, University of Arizona, Tucson, Arizona, USA

Abstract

This study reports the hybridization kinetics of double-stranded DNA probes for rapid molecular analysis. Molecular binding schemes based on double-stranded DNA probes have been developed for quantitative detection of various biomolecules, such as nucleic acids and DNA binding proteins recently. The thermodynamic competition between the target and the competitor in binding to the probe provides a highly specific mechanism for molecular detection. The kinetics of the double-stranded DNA probe, on the other hand, represent another key aspect toward its general applicability for a wide set of biomedical applications. Herein we report a systematic investigation of the kinetics of double-stranded DNA probes. The signal-to-background ratio and assay time of the double-stranded DNA probes are optimized at a high ionic strength (over 100 mM NaCl). Both the donor probe and the quencher probe sequences are shown to be important in the hybridization kinetics. A long sticky end of the probe is able to dramatically accelerate the kinetics of the assay. To provide a quantitative description of the kinetics, a two-stage binding model is developed to describe the major features of the kinetics of the assay. The sensitivity of the kinetic model and the dominant affinity constants are studied. The study provides a general guideline for the design of the probes for reducing the total assay time. With an appropriate design of the probes, the assay can be finished within minutes at room temperature.

Graphical abstract: Hybridization kinetics of double-stranded DNA probes for rapid molecular analysis

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Article information

DOI
https://doi.org/10.1039/B906077D
Article type
Paper
Submitted
26 Mar 2009
Accepted
06 May 2009
First published
22 May 2009

Analyst, 2009,134, 1675-1681

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Hybridization kinetics of double-stranded DNA probes for rapid molecular analysis

V. Gidwani, R. Riahi, D. D. Zhang and P. K. Wong, Analyst, 2009, 134, 1675 DOI: 10.1039/B906077D

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