Abstract
Nucleic acids have special ability to organize themselves into various non-canonical structures, including a four-stranded DNA structure termed G-quadruplex (G4) that has been utilized for diagnostic and therapeutic applications. Herein, we report the ability of G4 to distinguish dengue virus (DENV) based on its serotypes (DENV-1, DENV-2, DENV-3 and DENV-4) using a split G4-hemin DNAzyme configuration. In this system, two separate G-rich oligonucleotides are brought together upon target DNA strand hybridization to form a three-way junction architecture, allowing the formation of a G4 structure. The G4 formation in complexation with hemin can thus provide a signal readout by generating a DNAzyme that is able to catalyze H2O2-mediated oxidation of 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS). This results in a change of color providing a sensing platform for the colorimetric detection of DENV. In our approach, betaine and dimethyl sulfoxide were utilized for better G4 generation by enhancing the target-probe hybridization. In addition to this serotype-specific assay, a multi-probe cocktail assay, which is an all-in-one assay was also examined for DENV detection. The system highlights the potential of split G-quadruplex configurations for the development of DNA-based detection and serotyping systems in the future.
Highlights
Serving as a genetic information storage system for all living organisms as well as many viruses, deoxyribonucleic acid (DNA) has allowed scientists to identify key organisms based on sequences that are unique to speci c organism.[1]
We demonstrated a simple DNA-based system for dengue virus (DENV) serotyping and detection which takes advantage of DNAzymes generated from split G4-hemin complexes
This formation originated from single-stranded DNA molecules that can recognize and anneal to their complementary strands in a sample with stable hydrogen bond formation
Summary
The versatility of G4 has captured growing attention from different research groups for diagnostic assay development.[4,5,6,7] This has helped shape the way we perceive DNA structures today and has allowed researchers to construct functional structures, expanding the presumed traditional roles of DNA. Visually, as the base-speci c steric con guration of G4 allows the stacked G-quartets to bind to it In this way, binding regulates an enzymatic-like function that mimics peroxidase activity, generating a DNAzyme that catalyses H2O2-oxidation of 2,20-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS2À) to ABTS·-. The proposed assay utilizes split G4 structures as a reporter system for dengue DNA sensing The capability of this approach to distinguish different sequences which share low degrees of variation was tested. We demonstrated a simple DNA-based system for DENV serotyping and detection which takes advantage of DNAzymes generated from split G4-hemin complexes This formation originated from single-stranded DNA (ssDNA) molecules that can recognize and anneal to their complementary strands in a sample with stable hydrogen bond formation.
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