Advancements and Prospects in DNA-Based Bioanalytical Technology for Environmental Toxicant Detection

Abstract

The mounting global crisis of environmental pollution necessitates transformative advances in analytical technologies that combine speed, precision, and field applicability. To meet this demand, next-generation analytical platforms must achieve seamless integration of two critical features: molecular-level recognition fidelity and reliable signal transduction. DNA nanotechnology leverages sequence-specific molecular recognition and programmable self-assembly to enable both natural (e.g., riboswitches) and synthetic (e.g., aptamers, DNAzymes) biosensing modalities. The structural programmability and predictable Watson-Crick base pairing of DNA provide a modular framework for designing next-generation biosensors with tunable specificity and sensitivity. When integrated with portable point-of-care (POC) platforms, these biosensing systems enable field-deployable, rapid, and operator-agnostic detection of toxicants across diverse matrixes, making them highly suitable for complex environmental monitoring tasks. This perspective highlights the potential and strategic approaches for constructing biosensors utilizing DNA-based recognition elements and structural materials. It explores the progress in field-deployable DNA-based biosensors, which are revolutionizing the on-site detection of environmental toxicants. We also discuss the current challenges and future perspectives for DNA-based biosensing systems in environmental pollution monitoring, offering insights into their broader applications.

Keywords: DNA-based biosensing system; Recognition element; cell-free biosensor; environmental monitoring; field deployment.

1. Principles and Applications of DNA-Based Biosensing Systems

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Typical DNA-based recognition elements and strategies for sensing. The presence of an analyte triggers the allosteric effect of a natural recognition element, including riboswitch (A) and allosteric transcription factor (B), leading to the activation or inhibition of gene expression. Additionally, artificial recognition elements such as aptamer (C), DNAzyme (D), and G-quadruplex (E) can fold into conformationally specific structures that enable high-affinity binding to targets. SD: sensing domain of riboswitch, ED: expression effector domain of riboswitch, aTF: allosteric transcription factor, DBD: DNA-binding domain of aTF, EBD: effector-binding domain of aTF.

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Cell-free biosensors. (A) The system typically comprises three programmable modules: a highly synthetic DNA template, a recognition module, and a reporting module. These modules collectively regulate cell-free expression, where the target induces allosteric activation of the recognition element, performing signal reporting system functions facilitated by cell-free expression (CFE) substrates and ultimately generating quantifiable optical or catalytic signals proportional to target concentration. (B) Cell-free biosensor signal reporting pathways and system optimization. Cell-free biosensor signal reporting workflow progresses through three evolutionary stages: in vitro transcription-translation system (TXTL), in vitro transcription (IVT), and ligand-responsive artificial protein–protein communication (LIRAC), operationally achieving system simplification, accelerated response, and robustness enhancement. CRISPR/Cas: clustered regularly interspaced short palindromic repeat and CRISPR-associated protein. Portions of this figure were created with http://BioRender.com.

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DNA-based bioanalytical sensors. DNA recognition elements, such as aptamer, DNAzyme, or G4, sense the presence of target toxicants and activate the reactions in the dynamic signal transduction module, including conformational switching or toehold-mediated strand-displacement reaction (TMSDR). Finally, the reporting module generates detectable signals in various forms, such as fluorimetry, electrochemistry, colorimetry, or surface-enhanced Raman scattering (SERS), indicating the target toxicant concentration.

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DNA-based nanosensors. Based on DNA-based bioanalytical sensors, programmable DNA nanostructures have been utilized and integrated to engineer DNA nanosensors, serving multiple functions within the sensing system. These architectures play various critical roles within sensing systems, serving as structural scaffolds to immobilize elements, embedding recognition modules within nanostructured backbones to regulate spatially programmed assembly and functioning as dynamic nanomachines that facilitate signal transduction and amplification. DNA nanostructures have emerged as powerful tools to address several challenges faced by DNA-based sensors and presented a transformative approach by immobilizing functional elements within spatially confined architectures. This spatial restriction achieves focal amplification, enables directional reaction pathways, and facilitates optimized signal transduction and amplification mechanisms, collectively enhancing both sensing fidelity and dynamic functionality.

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Microfluidic chips. Microfluidic chips integrate sample mixing, reaction, incubation, and sensing within a single platform. (A) Detection of multiple metal ions on a single platform. Adapted with permission from ref . Copyright 2024 Elsevier. (B) Dual-mode output signals commonly enhance the reliability of detection results. Adapted with permission from ref . Copyright 2024 Elsevier. (C) Simultaneous detection of various pollutants. Adapted from ref . Copyright 2019 American Chemical Society. Portions of this figure were created with http://BioRender.com.

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Paper-based platform. A paper-based platform enables the rapid visual detection of toxicants. (A) Recognition of toxicants is translated into CRISPR-Cas system activation, allowing rapid detection using a commercial lateral flow assay. Adapted from ref . Copyright 2022 American Chemical Society. (B) Toxicant levels are converted into nucleic acid signals, which can be easily integrated with amplification techniques such as RPA to quantify the output signal via lateral flow easily. Adapted from ref . Available under a CC-BY 4.0 license. Copyright 2018 The Authors. (C) A lyophilized paper-based sensor, encapsulated using freeze-drying technology, can be rehydrated on-site for rapid environmental pollution testing. Adapted with permission from ref . Copyright 2022 Elsevier. Portions of this figure were created with http://BioRender.com.

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Portable miniaturized device. (A) Molecular events are represented as glucose levels and quantified with commercially available glucose monitors. Adapted from ref . Available under a CC-BY 4.0 license. Copyright 2021 The Authors. (B) Optical signals from ROSALID sensors are visualized with a 3D-printed hand-held illuminator. Adapted with permission from ref . Copyright 2020 Springer Nature. (C) Fluorescent signals from the LIRAC biosensing system are rapidly quantified with a portable detection device. Adapted with permission from ref . Copyright 2025 Wiley-VCH. Portions of this figure were created with http://BioRender.com.