A general approach for rational design of fluorescent DNA aptazyme sensors based on target-induced unfolding of DNA hairpins

Zhou, Zhaojuan; Xiao, Lü; Xiang, Yu; Zhou, Jun; Tong, Aijun

doi:10.1016/j.aca.2015.06.036

Cited by 14 publications

(10 citation statements)

References 66 publications

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“…Nevertheless, taken altogether, these results revealed that the developed bioassay can be successfully performed both in singleplex and multiplex formats with sensitivities in the nM range. This is in accordance with similar reported LOD values for other aptazymes (including those with a different catalytic cores), ranging from 6.9 pM to 18 μM when using fluorescencebased read-out [25,45,46].…”

Section: Dna-only Multiplex Bioassay For Detection Of Thrombin and Na Target Or Two Na Targetssupporting

confidence: 92%

DNA-only bioassay for simultaneous detection of proteins and nucleic acids

et al. 2021

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Graphical abstract Testing multiple biomarkers, as opposed to one, has become a preferred approach for diagnosing many heterogeneous diseases, such as cancer and infectious diseases. However, numerous technologies, including gold standard ELISA and PCR, can detect only one type of biomarker, either protein or nucleic acid (NA), respectively. In this work, we report for the first time simultaneous detection of proteins and NAs in the same solution, using solely functional NA (FNA) molecules. In particular, we combined the thrombin binding aptamer (TBA) and the 10-23 RNA-cleaving DNA enzyme (DNAzyme) in a single aptazyme molecule (Aptazyme 1.15-3′ ), followed by extensive optimization of buffer composition, sequences and component ratios, to establish a competitive bioassay. Subsequently, to establish a multiplex bioassay, we designed a new aptazyme (Aptazyme 2.20-5′ ) by replacing the target recognition and substrate sequences within Aptazyme 1.15-3′ . This designing process included an in silico study, revealing the impact of the target recognition sequence on the aptazyme secondary structure and its catalytic activity. After proving the functionality of the new aptazyme in a singleplex bioassay, we demonstrated the capability of the two aptazymes to simultaneously detect thrombin and NA target, or two NA targets in a multiplex bioassay. High specificity in target detection was achieved with the limits of detection in the low nanomolar range, comparable to the singleplex bioassays. The presented results deepen the barely explored features of FNA for diagnosing multiple targets of different origins, adding an extra functionality to their catalogue. Supplementary Information The online version contains supplementary material available at 10.1007/s00216-021-03458-6.

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Section: Dna-only Multiplex Bioassay For Detection Of Thrombin and Na Target Or Two Na Targetssupporting

confidence: 92%

DNA-only bioassay for simultaneous detection of proteins and nucleic acids

et al. 2021

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“…Based on this strategy, aptazymes are able to directly transduce molecular recognition into a quantitative catalytic event. Taking advantages of highly specific recognition property of aptamers and signal amplification property of NAEs via multiple‐turnover reactions, aptazymes have demonstrated their great promise in many in vitro applications …”

Section: Molecular Engineering Of Aptazyme‐based Nanomaterials Towardmentioning

confidence: 99%

Molecular Engineering of Functional Nucleic Acid Nanomaterials toward In Vivo Applications

Zhang

Lan

2019

Adv Healthcare Materials

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equally exciting and perhaps more challenging field of application is toward their in vivo applications, including living animals and human bodies, in diverse areas, [2] such as sensing, drug delivery, medical imaging, and cancer therapy. However, most of these nanomaterials lack selectivity toward targets of interests. Therefore, to employ these nanomaterials for a wider range of in vivo applications, various molecular engineering approaches have been employed to obtained nanomaterials functionalized with biomolecules to enable selective targeting. [3][4][5] Among these biomolecules, functional nucleic acids, or FNAs, have shown the most promise.FNAs are DNA or RNA molecules that can interact with or bind to a specific analyte, resulting in a conformational change and/or a catalytic reaction. [6] As their names suggested, ribozymes and deoxyribozymes (called DNAzyme hereafter) can act as enzymes in the presence of cofactors to catalyze various reactions, including cleavage and ligation of RNA/DNA, and phosphorylation and peroxidation of DNA. On the other hand, riboswitches and DNA aptamers are antibody-like receptors that can bind target molecules with high affinity and selectivity. Furthermore, the binding abilities of riboswitches and aptamers can be combined with the enzymatic function of ribozymes or DNAzymes to form aptazymes so that the binding of targets can inhibit or enhance the catalytic activities. These nucleic acids, including DNAzymes, aptamers, and aptazymes, are collectively known as FNAs to emphasize their functions beyond the traditional genetic storage and transformations roles for DNA and RNA. Unlike DNA and RNA inside biological systems, FNAs are isolated via a combinatorial technique known as in vitro selection, or a process also known as systematic evolution of ligands by exponential enrichment (SELEX). The discovery of new functions of nucleic acids has not only resulted in major advances in the fields of molecular biology and biochemistry, but also revolutionized sensors designs for both in vitro and in vivo applications. [7,8] Over the past decade, numerous FNA-based nanomaterials have been developed. [9][10][11][12] Among these FNA-based nanosystems, the functions of FNAs can be categorized into two types: diagnostic function and therapeutic function. For the diagnostic function, the FNAs serve either as the biorecognition ligands for direct sensing and imaging of the Recent advances in nanotechnology and engineering have generated many nanomaterials with unique physical and chemical properties. Over the past decade, numerous nanomaterials are introduced into many research areas, such as sensors for environmental monitoring, food safety, point-ofcare diagnostics, and as transducers for solar energy transfer. Meanwhile, functional nucleic acids (FNAs), including nucleic acid enzymes, aptamers, and aptazymes, have attracted major attention from the biomedical community due to their unique target recognition and catalytic properties. Benefiting from the recent progress of molecular engine...

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“…So far, however, the available examples of DNA aptazyme sensors are still limited in utilizing only several DNAzymes and DNA aptamers, most likely due to the lack of a general and simple approach for rational design. Zhou et al showed a general approach for designing fluorescent DNA aptazyme sensors [89]. Specifically, aptamers and DNAzymes are connected at the ends to avoid any change in their original sequences, therefore enabling the general use of different aptamers and DNAzymes in the design.…”

Section: Allosterically Regulated Dnazymes and Aptazymesmentioning

confidence: 99%

“…Many other strategies have been proposed to achieve an enhanced amplification using DNAzyme as a catalytic signal amplifier to construct biosensors [97]. Despite significant efforts in developing allosteric aptazyme-based sensors [98,99], only few sensors have been tested in biological samples [89,100,101]. This is probably due to the difficulty to transfer aptazymes developed for test tube to work in complex biological matrices.…”

Section: Allosterically Regulated Dnazymes and Aptazymesmentioning

confidence: 99%

Allosterically regulated DNA-based switches: From design to bioanalytical applications

Rossetti

Porchetta

2018

Analytica Chimica Acta

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DNA-based switches are structure-switching biomolecules widely employed in different bioanalytical applications. Of particular interest are DNA-based switches whose activity is regulated through the use of allostery. Allostery is a naturally occurring mechanism in which ligand binding induces the modulation and fine control of a connected biomolecule function as a consequence of changes in concentration of the effector. Through this general mechanism, many different allosteric DNA-based switches able to respond in a highly controlled way at the presence of a specific molecular effector have been engineered. Here, we discuss how to design allosterically regulated DNA-based switches and their applications in the field of molecular sensing, diagnostic and drug release.

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A general approach for rational design of fluorescent DNA aptazyme sensors based on target-induced unfolding of DNA hairpins

Cited by 14 publications

References 66 publications

DNA-only bioassay for simultaneous detection of proteins and nucleic acids

DNA-only bioassay for simultaneous detection of proteins and nucleic acids

Molecular Engineering of Functional Nucleic Acid Nanomaterials toward In Vivo Applications

Allosterically regulated DNA-based switches: From design to bioanalytical applications

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