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MicroRNAs (miRNAs) play a significant role in regulating diverse physiological processes, and are regarded as novel diagnostic biomarkers. However, the sensitive and reliable miRNA detection remains a huge challenge. Herein, we propose a proximity ligated initiated magnesium ion (Mg 2+ )-dependent DNAzyme-powered signal cascade for sensitive, accurate and reliable detection of miRNAs. Three signal amplification processes are involved in this approach, including the target miRNA recycle, DNAzyme powered substrate cleavage, and catalytic hairpin reaction (CHA). Based on this, the approach shows a low limit of detection of 523 aM and a wide detection range of 7 orders of magnitudes, which is comparable or superior to most of the former miRNA detection methods. In addition, the approach also possesses a high selectivity to target miRNA, suggesting a potential promising future of the approach for rapid detection of miRNAs in the application of developing novel tools for skin cancer diagnosis, and recovery evaluation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Cuproptosis, a kind of newly recognized cell death modality, shows enormous prospect in cancer treatment. The inducer of cuproptosis has more advantages in tumor therapy, especially that can trigger cuproptosis and chemodynamic therapy (CDT) simultaneously. However, cuproptosis is restricted to the deficiency of intracellular copper ions and the nonspecific delivery of copper-based ionophores. Therefore, high level delivery, responsive release, and utilizing synergistic-function of inducer become the key on cuproptosis-based oncotherapy. In this work, a cascade nanosystem is constructed for enhanced cuproptosis and CDT. In the weak acidic environment of tumor cells, DNA, zinc ions, and Cu + can release from the nanosystem. Since Cu + having superior performance in mediating both Fenton-like reaction and cuproptosis, the released Cu + induces cuproptosis and CDT efficiently, accompanied by Cu 2+ generation. Then Cu 2+ can be converted into Cu + partially by glutathione (GSH) to from a Cu + supply loop and ensure the synergistic action. Meanwhile, the consumption of GSH also contributes to cuproptosis and CDT in return. Finally, DNA and Zn 2+ form DNAzyme to shear catalase-related RNA, resulting in the accumulation of hydrogen peroxide and further enhancing combination therapy. These results provide a promising nanotherapeutic platform and may inspire the design for potential cancer treatment based on cuproptosis., (© 2023 Wiley-VCH GmbH.)

Multicomponent deoxyribozymes (MNAzymes) have great potential in gene therapy, but their ability to recognize disease tissue and further achieve synergistic gene regulation has rarely been studied. Herein, Arginylglycylaspartic acid (RGD)-modified Distearyl acylphosphatidyl ethanolamine (DSPE)-polyethylene glycol (PEG) (DSPE-PEG-RGD) micelle is prepared with a DSPE hydrophobic core to load the photothermal therapy (PTT) dye IR780 and the calcium efflux pump inhibitor curcumin. Then, the MNAzyme is distributed into the hydrophilic PEG layer and sealed with calcium phosphate through biomineralization. Moreover, RGD is attached to the outer tail of PEG for tumor targeting. The constructed nanomachine can release MNAzyme and the cofactor Ca 2+ under acidic conditions and self-assemble into an active mode to cleave heat shock protein (HSP) mRNA by consuming the oncogene miRNA-21. Silencing miRNA-21 enhances the expression of the tumor suppressor gene PTEN, leading to PTT sensitization. Meanwhile, curcumin maintains high intracellular Ca 2+ to further suppress HSP-chaperone ATP by disrupting mitochondrial Ca 2+ homeostasis. Therefore, pancreatic cancer is triple-sensitized to IR780-mediated PTT. The in vitro and in vivo results show that the MNAzyme-based nanomachine can strongly regulate HSP and PTEN expression and lead to significant pancreatic tumor inhibition under laser irradiation., (© 2023. The Author(s).)

Various binding modes of tunable metal organic frameworks (MOFs) and functional DNAzymes (Dzs) synergistically catalyze the emergence of abundant functional nanoplatforms. Given their serial variability in formation, structural designability, and functional controllability, Dzs@MOFs tend to be excellent building blocks for the precise "intelligent" manufacture of functional materials. To present a clear outline of this new field, this review systematically summarizes the progress of Dz integration into MOFs (MOFs@Dzs) through different methods, including various surface infiltration, pore encapsulation, covalent binding, and biomimetic mineralization methods. Atomic-level and time-resolved catalytic mechanisms for biosensing and imaging are made possible by the complex interplay of the distinct molecular structure of Dzs@MOF, conformational flexibility, and dynamic regulation of metal ions. Exploiting the precision of DNAzymes, MOFs@Dzs constructed a combined nanotherapy platform to guide intracellular drug synthesis, photodynamic therapy, catalytic therapy, and immunotherapy to enhance gene therapy in different ways, solving the problems of intracellular delivery inefficiency and insufficient supply of cofactors. MOFs@Dzs nanostructures have become excellent candidates for biosensing, bioimaging, amplification delivery, and targeted cancer gene therapy while emphasizing major advancements and seminal endeavors in the fields of biosensing (nucleic acid, protein, enzyme activity, small molecules, and cancer cells), biological imaging, and targeted cancer gene delivery and gene therapy. Overall, based on the results demonstrated to date, we discuss the challenges that the emerging MOFs@Dzs might encounter in practical future applications and briefly look forward to their bright prospects in other fields.

Cadmium is a heavy metal that is exceedingly hazardous to humans and can enter the body through tainted food or drink, causing severe harm. It is critical to develop a technology for detecting cadmium in food and water that is sensitive and accurate. One such approach, which employs nucleases, is uncommon. A cadmium(II) turn-on biosensor was successfully created in this work using repetitive cleavage of certain specific nucleases for signal conversion and sophisticated stem-loop qPCR (quantitative polymerase chain reaction) for quick signal amplification and output. The method has strong selectivity and sensitivity for precise quantification, with a detection limit of 6 nmol L -1 , i.e. 0.948 g L -1 , which is far lower than the 5.0 g L -1 set by the United States Environmental Protection Agency, and it also operates well in retail rice samples., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Panax ginseng and Panax quinquefolium are two valuable Chinese herbal medicines that should not be mixed because they differ in drug properties and efficacy. The traditional identification method is easily affected by subjective factors and cannot effectively distinguish between ginseng products. This study aimed to develop a new chemical analysis method to visually identify P. ginseng and P. quinquefolium. In this method, a large number of sequences containing G-quadruplex were generated by loop-mediated isothermal amplification, and the combination of G-quadruplex and hemin was used to form deoxyribozyme, which catalyzed the color change of H 2 O 2 . Artificial simulation of adulteration experiments revealed that this method could detect more than 20% adulterated P. quinquefolium. Compared with the traditional identification methods, this technology was simpler and more efficient, providing a reference for developing rapid visual identification methods and reagents for P. ginseng and P. quinquefolium., Competing Interests: Declaration of competing interest All authors disclosed no relevant relationships., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Pathogenic bacteria have a significant impact on food safety. Herein, an innovative dual-mode ratiometric aptasensor was constructed for ultrasensitive and accurate detection of Staphylococcus aureus (S. aureus) based on recycling of DNAzyme activation on gold nanoparticles-functionalized MXene nanomaterials (MXene@Au NPs). Electrochemiluminescent (ECL) emitter-labeled probe DNA (probe 2-Ru) containing the blocked DNAzyme was partly hybridized with aptamer and then captured by electrochemical (EC) indicator-labeled probe DNA (probe 1-MB) on electrode surface. When S. aureus presented, the conformation vibration of probe 2-Ru activated the blocked DNAzymes, leading to recycling cleavage of probe 1-MB and ECL tag close to electrode surface. Based on the reverse change tendencies of ECL and EC signals, aptasensor achieved S. aureus quantification from 5 to 10 8 CFU/mL. Moreover, the self-calibration characteristic of the aptasensor with dual-mode ratiometric readout ensured the reliable measurement of S. aureus in real samples. This work showed useful insight into sensing foodborne pathogenic bacteria., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Given the targeted binding ability and cleavage activity of the emerging CRISPR/Cas12a assay which transduces the target into its cleavage activity exhibited broadly prospective applications in integrated sensing and actuating system. Here, we elaborated a universal approach to quickly activate CRISPR/Cas12a for low-abundance biomarker detection based on the amplification strategy of a target-induced spherical nucleic acid enzyme (SNAzyme) network that could accelerate the output of activators. Specifically, multifunctional Y-shaped probes and hairpin probes (HPs, which contained the specific sequence of the activators of CRISPR/Cas12a and the substrate chain of DNAzyme) were rationally designed to construct SNAzyme. Target recognition induced disassembly of the Y-shaped probes, which released DNAzyme strands to active DNAzyme and accompanied by SNAzyme self-assembly into SNAzyme network. Interestingly, compared with randomly dispersed SNAzyme, the reaction kinetics of the SNAzyme network enhanced 1.6 times in response to Α-methyl acyl-CoA racemase (AMACR, a biomarker for prostate cancer), which was attributed to the promoted catalytic efficiency of DNAzyme by the confined SNAzyme network. Benefiting from these, the prepared biosensor based on electrochemiluminescence (ECL) platform by loading AuAg nanoclusters (AuAgNCs) into metal-organic framework-5 (MOF-5) exhibited satisfying detection performance for AMACR with a wide linear range (0.001 μg/mL to 100 μg/mL) and a low detection limit (1.0 × 10 -4  μg/mL, which exhibited significant potential in clinical diagnoses., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Dynamic monitoring of environmental Pb 2+ is of utmost importance for food safety and personal well-being. Herein, we report a novel, rapid, and practical fluorescence detection platform for Pb 2+ . The platform comprises two essential components: an engineered DNAzyme probe (EDP) and a responsive functionalized probe (RFP). The EDP demonstrates specific recognition of Pb 2+ and the subsequent release of free DNA fragments. The released DNA fragments are then captured using the RFP to form DNA complexes, which undergo multiple cascade amplification reactions involving polymerases and nickases, resulting in the generation of a large number of fluorescence signals. These signals can detect Pb 2+ at concentrations as low as 0.114 nmol/L, with a dynamic range spanning from 0.1 nmol/L to 50 nmol/L. Moreover, the platform exhibits excellent sensitivity and selectivity for Pb 2+ detection. To further validate its effectiveness, we successfully quantitatively detected lead contamination in water from Chaohu Lake, and the results aligned closely with those obtained using inductively coupled plasma-mass spectrometry (ICP-MS). Moreover, this platform is suitable for detecting Pb 2+ in seawater, soil, and fish samples. These findings confirm the suitability of the current detection platform for the dynamic assessment of Pb contamination in ecological environments, thereby contributing to environmental and food safety., Competing Interests: Declaration of Competing Interest The authors declare no competing financial interest., (Copyright © 2023 Elsevier B.V. All rights reserved.)

In this work, the orderly aggregated catalytic hairpin assembly (OA-CHA) was developed for synchronous ultrasensitive detection and high-efficiency colocalization imaging of dual-miRNAs by a carefully designed tetrahedral conjugated ladder DNA structure (TCLDS). Exactly, two diverse hairpin probes were fixed on tetrahedron conjugated DNA nanowires to form the TCLDS without fluorescence response, which triggered OA-CHA in the aid of output DNA 1 and output DNA 2 produced by targets miRNA-217 and miRNA-196a cycle to generate TCLDS with remarkable fluorescence response. Impressively, compared with the traditional CHA strategy, OA-CHA avoided the fluorescence group and quenching group from approaching again because of the spatial confinement effect to significantly enhance the fluorescence signal, resulting in the simultaneous ultrasensitive detection of dual-miRNAs with detection limits of 21 and 32 fM for miRNA-217 and miRNA-196a, respectively. Meanwhile, the TCLDS with lower diffusivity could achieve accurate localization imaging for reflecting the spatial distribution of dual-miRNAs in living cells. The strategy based on OA-CHA provided a flexible and programmable nucleic amplification method for the synchronous ultrasensitive detection and precise imaging of multiple biomarkers and had potential in disease diagnostics..

Herein, an ultrasensitive electrochemical biosensor for microRNA-155 (miR-155) detection based on the powerful catalytic and continuous walking signal amplification capability of 3D DNAzyme walker and the gold nanoparticles/graphene aerogels carbon fiber paper-based (AuNPs/GAs/CFP) flexible sensing electrode with excellent electrochemical performance was successfully constructed. In a proof-of-concept experiment, in the presence of miR-155, the DNAzyme strands anchored on the streptavidin-modified magnetic beads (MBs) silenced by locked strands can be activated, thus generating the walking arm of the 3D DNAzyme walker. Meanwhile, the substrate strands modified with Fe-MOF-NH 2 nanoparticles were evenly distributed on the surface of MBs and served as tracks of the 3D DNAzyme walker. Once the DNAzyme strand was activated, the catalytic site in the substrate strand can be cleaved in the presence of Mn 2+ , and a large number of stumps modified with Fe-MOF-NH 2 nanoparticles (output@Fe-MOF-NH 2 ) will be generated during the continuous and efficient walking cleavage of the DNAzyme walker, driving the recognition-catalysis-release cycle process for signal amplification. Immediately afterwards, the signal was read out through the base complementary pairing of capture probe (PS) immobilized on the surface of the paper-based flexible sensing electrode AuNPs/GAs/CFP and signal probes output@Fe-MOF-NH 2 , thus achieving the quantitative detection of miR-155. Under optimal experimental conditions, the designed 3D DNAzyme walker-based biosensor exhibited a relatively lower limit of detection (LOD) of 56.23 aM, with a linear range of 100 aM to 100 nM. Overall, the proposed 3D DNAzyme walker biosensor exhibited good interference and reproducibility, demonstrating a promising future in the field of clinical disease diagnosis., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Lead ion (Pb 2+ ) detection is critically important in environmental protection and health management. In this work, we developed a simple signal-enhanced colorimetric sensor for the detection of Pb 2+ based on the peroxidase-mimetic property of gold nanoparticles (AuNPs). When a certain concentration of Pb 2+ was added to a solution of DNAzyme-modified AuNPs, aggregation was triggered, and the result was an enhancement of the peroxidase-mimetic activity of AuNPs. Then, the chromogenic reaction of 3,3',5,5'-tetramethylbenzidine (TMB) by the catalyst of AuNPs was used for the sensitive UV-Vis and colorimetric detection of Pb 2+ . When a higher concentration of Pb 2+ was added, the greater amount of aggregation of AuNPs resulted in the enhancement of the UV-Vis adsorption of the solution at 652 nm, with a deepening of the blue color of the solution. After optimization of the experimental conditions, a linear relationship between the absorbance of oxidized TMB at 652 nm and the logarithm of Pb 2+ concentration was obtained, which had been divided into two parts (25 pM to 2.5 μM, and 2.5 μM to 250 μM). The detection limit was as low as 10 pM. The satisfactory specificity and rapid response of the sensor showed that it has promising application for the detection of Pb 2+ in real samples.

Glucose oxidase-loaded ZIF-90 metal-organic framework nanoparticles conjugated to hemin-G-quadruplexes act as functional bioreactor hybrids operating transient dissipative biocatalytic cascaded transformations consisting of the glucose-driven H 2 O 2 -mediated oxidation of Amplex-Red to resorufin or the glucose-driven generation of chemiluminescence by the H 2 O 2 -mediated oxidation of luminol. One system involves the fueled activation of a reaction module leading to the temporal formation and depletion of the bioreactor conjugate operating the nickase-guided transient biocatalytic cascades. The second system demonstrates the fueled activation of a reaction module yielding a bioreactor conjugate operating the exonuclease III-dictated transient operation of the two biocatalytic cascades. The temporal operations of the bioreactor circuits are accompanied by kinetic models and computational simulations enabling us to predict the dynamic behavior of the systems subjected to different auxiliary conditions.

Emulating native transient transcription machineries modulating temporal gene expression by synthetic circuits is a major challenge in the area of systems chemistry. Three different methods to operate transient transcription machineries and to modulate the gated transcription processes of target RNAs are introduced. One method involves the design of a reaction module consisting of transcription templates being triggered by promoter fuel strands transcribing target RNAs and in parallel generating functional DNAzymes in the transcription templates, modulating the dissipative depletion of the active templates and the transient operation of transcription circuits. The second approach involves the application of a reaction module consisting of two transcription templates being activated by a common fuel promoter strand. While one transcription template triggers the transcription of the target RNA, the second transcription template transcribes the anti-fuel strand, displacing the promoter strand associated with the transcription templates, leading to the depletion of the transcription templates and to the dynamic transient modulation of the transcription process. The third strategy involves the assembly of a reaction module consisting of a reaction template triggered by a fuel promoter strand transcribing the target RNA. The concomitant nickase-stimulated depletion of the promoter strand guides the transient modulation of the transcription process. Via integration of two parallel fuel-triggered transcription templates in the three transcription reaction modules and application of template-specific blocker units, the parallel and gated transiently modulated transcription of two different RNA aptamers is demonstrated. The nickase-stimulated transiently modulated transcription reaction module is applied as a functional circuit guiding the dynamic expression of gated, transiently operating, catalytic DNAzymes.

Sensitive and accurate detection of multiplex foodborne pathogens is crucial for food safety. In this work, a dual-mode and dual-target biosensor regulated by a Tesla valve was established for simultaneously determining Escherichia coli O157:H7 (E. coli) and Salmonella typhimurium (S. T). Two zeolitic imidazolate framework (ZIF-8) signal probes decorated with electroactive materials (ferrocene or methylene blue), DNAzyme, and different phages were synthesized to specifically recognize the targets and generate fluorescent/electrochemical dual-mode signals. In the presence of bacteria, they were captured and enriched on two individual working electrodes through the modified 4-mercaptophenylboric acid. The encoded signal probes added on different working electrodes could be conjugated with the corresponding target bacteria depending on the specificity of phages. Under the acidic condition, the DNAzyme could catalyze click chemistry for fluorescent signals. Simultaneously, the released ferrocene and methylene blue from ZIF-8 could generate electrochemical signals at different potentials. Benefiting from the flow regulation feature of the Tesla valve, the triggered fluorescent and electrochemical signals in the two individual electrodes would not influence each other, achieving simultaneous dual-mode and dual-target determination of foodborne pathogens. It depicted good linearity ranged 10-10 7  CFU mL -1 . And the corresponding detection of limits were 5 CFU mL -1 and 8 CFU mL -1 for two bacteria, respectively. A low false positive was realized through the dual-mode strategy. The proposed biosensor can not only on-site, specifically, and sensitively determine E. coli and S. T, but also provide the wide prospect in rapid screening of other foodborne pathogens., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Telomerase, as a specialized reverse transcriptase, plays a vital role in early cancer diagnostics and prognosis; thus, developing efficient sensing technologies is of vital importance. Herein, an innovative "signal-on-off" photoelectrochemical (PEC) sensing platform was developed for ultrasensitive evaluation of telomerase activity based on an electron-transfer tunneling distance regulation strategy and DNAzyme-triggerable biocatalytic precipitation. Concretely, cascade internal electric fields between CuInS 2 quantum dots (QDs), graphitic carbon nitride nanosheets (g-C 3 N 4 NSs), and TiO 2 nanorod arrays (NRAs) were developed to realize cascade electron extraction and hole transfer. Enabled by such a design, an effective "signal-on" state to gain a progressively enhanced PEC output was designed by suppressing the photogenerated electron-hole pair recombination. With the introduction of hairpin probe H2 and the subsequent extension of the primer sequence driven by the target telomerase, the CuInS 2 QDs labeled with hairpin probe H1 were programmatically unfolded, resulting in CuInS 2 QDs' close proximity to the working electrode away from the cascade interface, accompanied by the formation of G-quadruplex/hemin complexes. The gradual undermining of tunneling distance and implantation of DNAzyme-initiating biocatalytic precipitation tremendously induced the sluggish migration kinetics of the photoinduced charge, accompanied by the photocurrent intensity decrement, leading to the "signal-off" state. Under optimized conditions, the as-prepared PEC biosensor realizes ultrasensitive detection of telomerase activity from 10 to 10 5 cell·mL -1 with a detection limitation of 3 cells·mL -1 . As a proof of concept, this well-designed method provides new insights into signal amplification for telomerase activity evaluation and also presents promising potential for further development in drug screening, healthcare diagnostics, and biological assays.

Non-nucleic acid targets have posed a serious challenge to food safety. The detection of non-nucleic acid targets can enable us to monitor food contamination in a timely manner. In recent years, the CRISPR/Cas system has been extensively explored in biosensing. However, there is a lack of a summary of CRISPR/Cas-powered detection tailored to non-nucleic acid targets involved in food safety. This review comprehensively summarizes the recent advances on the construction of CRISPR/Cas-powered detection and the promising applications in the field of food safety related non-nucleic acid targets. The current challenges and futuristic perspectives are also proposed accordingly. The rapidly evolving CRISPR/Cas system has provided a powerful propellant for non-nucleic acid target detection via integration with aptamer and/or DNAzyme. Compared with traditional analytical methods, CRISPR/Cas-powered detection is conceptually novel, essentially eliminates the dependence on large instruments, and also demonstrates the capability for rapid, accurate, sensitive, and on-site testing.

Cereulide, the exotoxin of emetic Bacillus cereus , has garnered considerable attention due to its capacity to produce foodborne poisonings and great chemical stability. Herein, a G-quadruplex-hemin DNAzyme-based biosensor was developed to detect cereulide in the homogeneous solution. Due to the special ring structure and high affinity to K + , cereulide can be attracted and intercalated into the G-quadruplex; thus, the properties of the G4 DNAzyme can be altered. The melting temperature ( T m ) of the G4 DNAzyme in the presence or absence of cereulide was 58.75 and 50.10 °C, respectively, proving the intercalation of cereulide into the G4 DNAzyme. By using the polychromic fluorescence modality of CdTe quantum dots and o -phenylenediamine to assess the variation in the catalytic activity of the DNAzyme, the intercalation of cereulide had bidirectional effects in G4 DNAzyme-mediated reactions, showing that the fluorescence intensity of CdTe quantum dots displayed a linear relationship with the concentration of cereulide from 0.16 to 40 μg/mL with the limit of detection (LOD) of 0.10 μg/mL, while the fluorescence intensity of DAP exhibited a linear relationship with the concentration of cereulide from 0.02 to 40 μg/mL with the LOD of 0.01 μg/mL. It will be a perspective step of controlling cereulide as a hazardous material in food or the environment.

One of the highly attractive research directions in the electrochemiluminescence (ECL) field is how to regulate and improve ECL efficiency. Quantum dots (QDs) are highly promising ECL materials due to their adjustable luminescence size and strong luminous efficiency. MoS 2 NSs@QDs, an ECL emitter, is synthesized via hydrothermal methods, and its ECL mechanism is investigated using cyclic voltammetry and ECL-potential curves. Then, a stable and vertical attachment of a triplex DNA (tsDNA) probe to the MoS 2 nanosheets (NSs) is applied to the electrode. Next, an innovative ECL sensor is courageously empoldered for precise and ultrasensitive detection of target miRNA-199a through the agency of ECL-resonance energy transfer (RET) strategy and a dextrous target-initiated catalytic three-arm DNA junction assembly (CTDJA) based on a toehold strand displacement reaction (TSDR) signal amplification approach. Impressively, the ingenious system not only precisely regulates the distance between energy donor-acceptor pairs leave energy less loss and more ECL-RET efficiency, but also simplifies the operational procedure and verifies the feasibility of this self-assembly process without human intervention. This study can expand MoS 2 NSs@QDs utilization in ECL biosensing applications, and the proposed nucleic acid amplification strategy can become a miracle cure for ultrasensitive detecting diverse biomarkers, which helps researchers to better study the tumor mechanism, thereby unambiguously increasing cancer cure rates and reducing the risk of recurrence.

Due to the advantages of its numerous modification sites, predictable structure, high thermal stability, and excellent biocompatibility, DNA is the ideal choice as a key component of biosensors. DNA biosensors offer significant advantages over existing bioanalytical techniques, addressing limitations in sensitivity, selectivity, and limit of detection. Consequently, they have attracted significant attention from researchers worldwide. Here, we exemplify four foundational categories of functional nucleic acids: aptamers, DNAzymes, i-motifs, and G-quadruplexes, from the perspective of the structure-driven functionality in constructing DNA biosensors. Furthermore, we provide a concise overview of the design and detection mechanisms employed in these DNA biosensors. Noteworthy advantages of DNA as a sensor component, including its programmable structure, reaction predictility, exceptional specificity, excellent sensitivity, and thermal stability, are highlighted. These characteristics contribute to the efficacy and reliability of DNA biosensors. Despite their great potential, challenges remain for the successful application of DNA biosensors, spanning storage and detection conditions, as well as associated costs. To overcome these limitations, we propose potential strategies that can be implemented to solve these issues. By offering these insights, we aim to inspire subsequent researchers in related fields.

In the developed assay, multiorbital 3D DNA walker (MO DNA walker) was applied as signal amplified protocol for enhancing the detection signal of the photothermal biosensor, which was designed for sensitive detection of miRNA based on the H 2 S triggered conversation of photothermal reagent. When the target molecule is present, the DNA walking strand was released and then hybridize with track strands. The landing of walking particles (WPT) on the tracking particles (TPT) promotes the relative movement of the WPT around TPT, thus releasing large amount of horseradish peroxidase (HRP) with the aid of DNAzyme. After reacting with Na 2 S 2 O 3 and H 2 O 2 , multiple H 2 S can be generated in situ based on the catalytic ability of HRP. Meanwhile, cubic Prussian blue (CPB) was synthesized and exhibited superior photothermal response, which can be served as efficient photothermal reagent and H 2 S responsive acceptor. Significantly, the photothermal signal of CPB could be obviously reduced after challenged with H 2 S ascribed to synchronous reaction between the ferric ion (Fe 3+ ) and H 2 S. The improved walking area and freedom enable significant signal amplification, enhancing the biosensor's performance. Under ideal circumstances, the proposed photothermal assay demonstrated excellent performance for determination of miRNA-21., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Austria, part of Springer Nature.)

Multidrug resistance (MDR) is a major cause of chemotherapy failure in oncology, and gene therapy is an excellent measure to reverse MDR. However, conventional gene therapy only modulates the expression of MDR-associated proteins but hardly affects their existing function, thus limiting the efficiency of tumor treatment. Herein, we designed a photoactivated DNA nanodrug (MCD@TMPyP 4 @DOX) to improve tumor chemosensitivity through the downregulation of MDR-related genes and mitochondria-targeted photodynamic therapy (PDT). The self-assembled DNA nanodrug encodes the mucin 1 (MUC1) aptamer and the cytochrome C (CytC) aptamer to facilitate its selective targeting to the mitochondria in tumor cells; the encoded P-gp DNAzyme can specifically cleave the substrate and silence MDR1 mRNA with the help of Mg 2+ cofactors. Under near-infrared (NIR) light irradiation, PDT generates reactive oxygen species (ROS) that precisely damage the mitochondria of tumor cells and break single-stranded DNA (ssDNA) to activate MCD@TMPyP 4 @DOX self-disassembly for release of DOX and DNAzyme. We have demonstrated that this multifunctional DNA nanodrug has high drug delivery capacity and biosafety. It enables downregulation of P-gp expression while reducing the ATP on which P-gp pumps out drugs, improving the latency of gene therapy and synergistically reducing DOX efflux to sensitize tumor chemotherapy. We envision that this gene-modulating DNA nanodrug based on damaging mitochondria is expected to provide an important perspective for sensitizing tumor chemotherapy.

Highly sensitive and selective detection of microRNA-21 (miRNA-21) in biological samples is critical for the disease diagnosis and cancer treatment. In this study, a nitrogen-doped carbon dots (N-CDs)-based ratio fluorescence sensing strategy was constructed for miRNA-21 detection with high sensitivity and excellent specificity. Bright-blue N-CDs (λ ex /λ em  = 378 nm/460 nm) were synthesized by facile one-step microwave-assisted pyrolysis method by using uric acid as the single precursor, and the absolute fluorescence quantum yield and fluorescence lifetime of N-CDs were 35.8% and 5.54 ns separately. The padlock probe hybridized with miRNA-21 firstly and then was cyclized by T4 RNA ligase 2 to form a circular template. At the present of dNTPs and phi29 DNA polymerase, the oligonucleotide sequence in miRNA-21 was prolonged to hybridize with the surplus oligonucleotide sequences in circular template, generating long and reduplicated oligonucleotide sequences containing abundant guanine nucleotides. Separate G-quadruplex sequences were generated after the addition of Nt.BbvCI nicking endonuclease, and then hemin bound with G-quadruplex sequence to construct the G-quadruplex DNAzyme. Such G-quadruplex DNAzyme catalyzed the redox reaction of o-phenylenediamine (OPD) with H 2 O 2 , finally producing the yellowish-brown 2,3-diaminophenazine (DAP) (λ em  = 562 nm). Due to the inner filter effect between N-CDs and DAP, the ratio fluorescence signal of DAP with N-CDs was utilized for sensitive detection of miRNA-21 with detection limit of 0.87 pM. Such approach has practical feasibility and excellent specificity for miRNA-21 analysis during highly homological miRNA family in HeLa cell lysates and human serum samples., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Traditional hybridization chain reaction (HCR) as a popular isothermal amplification technique shows some inevitable disadvantages in bioanalysis due to its relatively slow kinetics, which could be markedly promoted when the HCR initiator occurs under tension. Herein, a poly DNA tetrahedrons (pTDNs)-mediated HCR was successfully constructed to make its initiator in a stretched state by long-range electrostatic forces owing to the superimposed electrostatic interactions derived from the synthesized pTDNs, and it was hypothesized that it could remarkably enhance HCR performance, which was testified by theoretical simulations and experimental studies. Consequently, pTDNs-mediated HCR was applied to develop a novel immunoassay for rapid and sensitive detection of aflatoxin B1 as a proof-of-concept, and its signal amplification was attributed to the increased G 4 DNAzyme that loaded on the second antibody. Our work paves a promising way using simple DNA frameworks alone to heighten HCR kinetics for reaction speed improvement and signal amplification in bioanalysis.

The increasing prevalence of antimicrobial resistance has called for improved diagnostic testing of pathogenic bacteria. However, the development of rapid, cost-effective, and easy-to-use tests for bacterial infections remains a constant challenge. Here, we report a class of modular hydrogel membrane carriers incorporated with composite DNAzymes, which enable rapid and highly sensitive detection of pathogenic bacteria gene target analytes. We apply free radical polymerization to incorporate composite DNAzymes, consisting of an RNA substrate component and a DNAzyme component (e.g., 10-23 or 8-17 DNAzymes), into polyethylene glycol diacrylate polymer networks. Initiated by a nucleic acid target acting as an assembly facilitator, multicomponent DNAzymes are combined to cleave the RNA substrate component in the hydrogel carriers, which releases the DNAzyme component to cleave RNA reporter probes to generate fluorescence. We modulate the morphology, composition, and microporous structures of the DNAzyme carriers to achieve quantitative assay performance. We demonstrate a rapid and high-sensitivity detection of C. trachomatis gene target analytes as low as 50 fM in a short assay time of 25 min. The work represents a crucial step forward in the development of a generic, isothermal, and protein enzyme-free pathogenic bacteria testing platform technology.

DNA-based enzymes, or DNAzymes, are single-stranded DNA sequences with the ability to catalyze various chemical reactions, including the cleavage of the bond between two RNA nucleotides. Lately, an increasing interest has been observed in these RNA-cleaving DNAzymes in the biosensing and therapeutic fields for signal generation and the modulation of gene expression, respectively. Additionally, multiple efforts have been made to study the effects of the reaction environment and the sequence of the catalytic core on the conversion of the substrate into product. However, most of these studies have only reported alterations of the general reaction course, but only a few have focused on how each individual reaction step is affected. In this work, we present for the first time a mathematical model that describes and predicts the reaction of the 10-23 RNA-cleaving DNAzyme. Furthermore, the model has been employed to study the effect of temperature, magnesium cations and shorter substrate-binding arms of the DNAzyme on the different kinetic rate constants, broadening the range of conditions in which the model can be exploited. In conclusion, this work depicts the prospects of such mathematical models to study and anticipate the course of a reaction given a particular environment.

Biological and clinical studies have indicated that aberrant expression of circMTO1 served as a crucial biomarker for the diagnosis and prognosis of hepatocellular carcinoma (HCC) patients as well as a potential therapeutic target. However, the detection of circRNAs currently faces challenges such as homologous linear RNA interference and low-expression abundance of certain circRNAs. Therefore, we developed a triple amplification method based on catalytic hairpin assembly (CHA) activation by back-splice junction (BSJ), resulting in CHA products that triggered primer exchange reaction to generate DNAzyme. Subsequently, DNAzyme cleaved the fluorescent reporter chain, enabling ultrasensitive detection of hepatocellular carcinoma-associated circMTO1 through the output fluorescence signal. The catalytic hairpin opening sequence within CHA specifically targeted the BSJ sequence in circRNA, thereby avoiding false positive signals observed in circRNA assays due to the recognition of homologous linear RNA molecules. Moreover, this triple amplification method facilitated sensitive detection of circRNA and addressed the issue of low-abundance expression levels associated with circMTO1 in HCC samples. Notably, our newly designed assay for detecting circRNA exhibited a linear range from 1 fM to 100 nM with a detection limit of 0.265 fM. Furthermore, it demonstrated excellent and consistent performance even within complex systems. Our proposed method enabled the specific and sensitive detection of circMTO1 in various cancer cells and blood samples from HCC patients, providing an innovative approach for investigating the role of circRNA in tumorigenesis and development while promoting its clinical application.

Since their first discovery in 1994, DNAzymes have been extensively applied in biosensing and therapy that act as recognition elements and signal generators with the outstanding properties of good stability, simple synthesis, and high sensitivity. One subset, RNA-cleaving DNAzymes, is widely employed for diverse applications, including as reporters capable of transmitting detectable signals. In this review, the recent advances of RNA-cleaving DNAzyme-based amplification strategies in scaled-up biosensing are focused, the application in diagnosis and disease treatment are also discussed. Two major types of RNA-cleaving DNAzyme-based amplification strategies are highlighted, namely direct response amplification strategies and combinational response amplification strategies. The direct response amplification strategies refer to those based on novel designed single-stranded DNAzyme, and the combinational response amplification strategies mainly include two-part assembled DNAzyme, cascade reactions, CHA/HCR/RCA, DNA walker, CRISPR-Cas12a and aptamer. Finally, the current status of DNAzymes, the challenges, and the prospects of DNAzyme-based biosensors are presented., (© 2023 Wiley-VCH GmbH.)

DNAzyme-based gene regulation shows great potential for the therapy of many cancers. However, ineffective delivery and insufficient cofactor supply pose challenges for potent gene therapy. In this study, we constructed a smart metal-polyphenol-DNAzyme nanoplatform (TA-Mn@Dz NPs) with intrinsic stability, effective delivery, and cofactor self-supply ability for gene-chemodynamic synergistic tumor therapy. Tannic acid, a plant-derived polyphenol, acts as an intermediate structural unit to mediate the assembly of Mn 2+ /DNAzyme and tumor acid environment-responsive nanocarriers. Intracellularly, the acidic environment triggers the decomposition of TA-Mn@Dz NPs to release DNAzyme and Mn 2+ . The Mn 2+ ion not only boosts the catalytic cleavage of surviving mRNA for effective gene therapy but also activates chemodynamic therapy (CDT), generating highly toxic ·OH from endogenous H 2 O 2 . When tail intravenously injected into MCF-7 tumor-bearing mice, the TA-Mn@Dz NPs display desirable synergistic gene-chemodynamic antitumor effects, paving the way for developing DNAzyme-based multifunctional theranostic platforms for biomedical applications. STATEMENT OF SIGNIFICANCE: 1. A smart metal-polyphenol-DNAzyme nanoplatform was constructed for gene-chemodynamic synergistic tumor therapy. 2. Tannic acid act as intermediate structural units to mediate the assembly of Mn 2+ /DNAzyme and tumor acid environment-responsive nanocarriers. 3. The Mn 2+ -ion could not only boost the catalytic cleavage of surviving mRNA for effective gene therapy, but also catalyze endogenous H 2 O 2 to form cytotoxic hydroxyl radicals for chemodynamic therapy. 4. Our work paves an extremely simple way to integrate gene therapy with CDT for the dual-catalytic tumor treatment., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)