A dual-cycling biosensor for target DNA detection based on the toehold-mediated strand displacement reaction and exonuclease III assisted amplification

Yu, Ling; Zhang, Xiao Fang; Chen, Xiao Hui; Liu, Li; Wang, Xiao Hu; Wang, De Shou; Li, Nian Bing; Luo, Hong Qun

doi:10.1039/c7nj05191c

Cited by 7 publications

(6 citation statements)

References 33 publications

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“…A considerable amount of experimental and theoretical work has been devoted to finding means to eliminate or mitigate against these reactions. ,,− Dynamic DNA nanotechnology has seen the greatest amount of activity in research directed to the practical application in medical diagnostics , and therapeutics, − an area that appears especially attractive due to the biocompatibility and the ease with which nucleic acid based devices and systems can be interfaced with biological DNA and RNA. DNA hybridization networks, functioning as nucleic acid detectors, achieving attomolar − and even subattomolar DNA detection sensitivity − have recently been reported.…”

Section: Introductionmentioning

confidence: 99%

Principles and Applications of Nucleic Acid Strand Displacement Reactions

2019

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Dynamic DNA nanotechnology, a subfield of DNA nanotechnology, is concerned with the study and application of nucleic acid strand-displacement reactions. Strand-displacement reactions generally proceed by three-way or four-way branch migration and initially were investigated for their relevance to genetic recombination. Through the use of toeholds, which are single-stranded segments of DNA to which an invader strand can bind to initiate branch migration, the rate with which strand displacement reactions proceed can be varied by more than 6 orders of magnitude. In addition, the use of toeholds enables the construction of enzyme-free DNA reaction networks exhibiting complex dynamical behavior. A demonstration of this was provided in the year 2000, in which strand displacement reactions were employed to drive a DNAbased nanomachine et al. Nature 2000, 406, 605−608). Since then, toeholdmediated strand displacement reactions have been used with ever increasing sophistication and the field of dynamic DNA nanotechnology has grown exponentially. Besides molecular machines, the field has produced enzyme-free catalytic systems, all DNA chemical oscillators and the most complex molecular computers yet devised. Enzyme-free catalytic systems can function as chemical amplifiers and as such have received considerable attention for sensing and detection applications in chemistry and medical diagnostics. Strand-displacement reactions have been combined with other enzymatically driven processes and have also been employed within living cells (Groves, B.; et al. Nat. Nanotechnol. 2015, 11, 287−294). Strand-displacement principles have also been applied in synthetic biology to enable artificial gene regulation and computation in bacteria. Given the enormous progress of dynamic DNA nanotechnology over the past years, the field now seems poised for practical application.

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Section: Introductionmentioning

confidence: 99%

Principles and Applications of Nucleic Acid Strand Displacement Reactions

2019

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“…With the advent of DNA nanotechnology, fluorescence resonance energy transfer (FRET)-based sensors have especially surged in popularity for their ability to detect a variety of targets including nucleic acids, proteins, and small molecules. Due to their simple and highly predictable design, ability to be regenerated through the toehold-mediated strand displacement (TMSD) process [4,5,6], and specificity towards pre-defined targets, DNA-based sensors have found applications in fluorescent hybridization assays [2,7,8], aptamer-based assays [7,9,10], electrochemical assays [3,11], surface enhanced Raman spectroscopy [12,13], and surface plasmon resonance analysis of biomolecules [14,15], just to name a few. Among these, bulk fluorescence methods are popular owing to their flexibility in sensor design, easy operation, and high sensitivity.…”

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

“…In order to achieve better detection limits, bulk FRET strategies are often coupled with signal amplification steps such as exonuclease-III (ExoIII)-assisted amplification, rolling circle amplification (RCA), or hybridization chain reaction (HCR), all of which introduce a significant complexity to the assays [8,9,18]. For example, an enzymatic amplification requires a multi-step process and use of enzyme, which is costly and sensitive to reaction conditions such as temperature, pH, and enzyme inhibitors [8,18]. Other emerging examples demonstrating a sensitive detection of DNA down to low picomolar and high attomolar concentrations in an amplification-free format involve complex systems such as quantum dots, nanoparticles, cationic polymers, or microarrays [19,20,21,22,23,24,25,26,27,28,29].…”

Section: Introductionmentioning

confidence: 99%

“…In addition to sensitive and quantitative detection, another highly desirable feature of nucleic acid sensors is the ability to distinguish a point mutation called single nucleotide polymorphisms (SNPs), as more and more nucleic acid sequences have been found as biomarkers [37,38,39,40,41,42]. The single nucleotide substitutions of a genome are the most abundant genetic polymorphisms and are regarded as genetic markers for identifying inherited diseases and also for the development of drug candidates [8,43,44]. In this regard, a variety of methods spanning from single molecule to bulk analysis have been developed to distinguish SNPs [31,45,46], for example, enzyme assisted allele-specific biochemical reactions, electrochemical techniques, selective ligation with a fluorescent-modified DNA, synthetic nanopores containing molecular probes, etc.…”

Section: Introductionmentioning

confidence: 99%

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Single-Step FRET-Based Detection of Femtomoles DNA

Sapkota

Kaur

Megalathan

et al. 2019

Sensors

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Sensitive detection of nucleic acids and identification of single nucleotide polymorphism (SNP) is crucial in diagnosis of genetic diseases. Many strategies have been developed for detection and analysis of DNA, including fluorescence, electrical, optical, and mechanical methods. Recent advances in fluorescence resonance energy transfer (FRET)-based sensing have provided a new avenue for sensitive and quantitative detection of various types of biomolecules in simple, rapid, and recyclable platforms. Here, we report single-step FRET-based DNA sensors designed to work via a toehold-mediated strand displacement (TMSD) process, leading to a distinct change in the FRET efficiency upon target binding. Using single-molecule FRET (smFRET), we show that these sensors can be regenerated in situ, and they allow detection of femtomoles DNA without the need for target amplification while still using a dramatically small sample size (fewer than three orders of magnitude compared to the typical sample size of bulk fluorescence). In addition, these single-molecule sensors exhibit a dynamic range of approximately two orders of magnitude. Using one of the sensors, we demonstrate that the single-base mismatch sequence can be discriminated from a fully matched DNA target, showing a high specificity of the method. These sensors with simple and recyclable design, sensitive detection of DNA, and the ability to discriminate single-base mismatch sequences may find applications in quantitative analysis of nucleic acid biomarkers.

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