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Biosensors Jun 2023DNA-mediated nanotechnology has become a research hot spot in recent decades and is widely used in the field of biosensing analysis due to its distinctive properties of... (Review)
Review
DNA-mediated nanotechnology has become a research hot spot in recent decades and is widely used in the field of biosensing analysis due to its distinctive properties of precise programmability, easy synthesis and high stability. Multi-mode analytical methods can provide sensitive, accurate and complementary analytical information by merging two or more detection techniques with higher analytical throughput and efficiency. Currently, the development of DNA-mediated multi-mode analytical methods by integrating DNA-mediated nanotechnology with multi-mode analytical methods has been proved to be an effective assay for greatly enhancing the selectivity, sensitivity and accuracy, as well as detection throughput, for complex biological analysis. In this paper, the recent progress in the preparation of typical DNA-mediated multi-mode probes is reviewed from the aspect of deoxyribozyme, aptamer, templated-DNA and G-quadruplex-mediated strategies. Then, the advances in DNA-mediated multi-mode analytical methods for biological samples are summarized in detail. Moreover, the corresponding current applications for biomarker analysis, bioimaging analysis and biological monitoring are introduced. Finally, a proper summary is given and future prospective trends are discussed, hopefully providing useful information to the readers in this research field.
Topics: Biosensing Techniques; DNA; Nanotechnology; DNA Probes; Oligonucleotides
PubMed: 37504092
DOI: 10.3390/bios13070693 -
Molecular Ecology Dec 2023Exhaustive biodiversity data, covering all the taxa in an environment, would be fundamental to understand how global changes influence organisms living at different... (Review)
Review
Exhaustive biodiversity data, covering all the taxa in an environment, would be fundamental to understand how global changes influence organisms living at different trophic levels, and to evaluate impacts on interspecific interactions. Molecular approaches such as DNA metabarcoding are boosting our ability to perform biodiversity inventories. Nevertheless, even though a few studies have recently attempted exhaustive reconstructions of communities, holistic assessments remain rare. The majority of metabarcoding studies published in the last years used just one or two markers and analysed a limited number of taxonomic groups. Here, we provide an overview of emerging approaches that can allow all-taxa biological inventories. Exhaustive biodiversity assessments can be attempted by combining a large number of specific primers, by exploiting the power of universal primers, or by combining specific and universal primers to obtain good information on key taxa while limiting the overlooked biodiversity. Multiplexes of primers, shotgun sequencing and capture enrichment may provide a better coverage of biodiversity compared to standard metabarcoding, but still require major methodological advances. Here, we identify the strengths and limitations of different approaches, and suggest new development lines that might improve broad scale biodiversity analyses in the near future. More holistic reconstructions of ecological communities can greatly increase the value of metabarcoding studies, improving understanding of the consequences of ongoing environmental changes on the multiple components of biodiversity.
Topics: Biodiversity; DNA; DNA Barcoding, Taxonomic; DNA Primers; Ecology
PubMed: 36762839
DOI: 10.1111/mec.16881 -
Proceedings of the National Academy of... Aug 2023Electronic detection of DNA oligomers offers the promise of rapid, miniaturized DNA analysis across various biotechnological applications. However, known all-electrical...
Electronic detection of DNA oligomers offers the promise of rapid, miniaturized DNA analysis across various biotechnological applications. However, known all-electrical methods, which solely rely on measuring electrical signals in transducers during probe-target DNA hybridization, are prone to nonspecific electrostatic and electrochemical interactions, subsequently limiting their specificity and detection limit. Here, we demonstrate a nanomechanoelectrical approach that delivers ultra-robust specificity and a 100-fold improvement in detection limit. We drive nanostructural DNA strands tethered to a graphene transistor to oscillate in an alternating electric field and show that the transistor-current spectra are characteristic and indicative of DNA hybridization. We find that the inherent difference in pliability between unpaired and paired DNA strands leads to the spectral characteristics with minimal influence from nonspecific electrostatic and electrochemical interactions, resulting in high selectivity and sensitivity. Our results highlight the potential of high-performance DNA analysis based on miniaturized all-electronic settings.
Topics: DNA; Nucleic Acid Hybridization; DNA Probes; Graphite; Hybridization, Genetic; Biosensing Techniques
PubMed: 37549255
DOI: 10.1073/pnas.2306130120 -
Nature Communications Jul 2023The T4 bacteriophage gp41 helicase and gp61 primase assemble into a primosome to couple DNA unwinding with RNA primer synthesis for DNA replication. How the primosome is...
The T4 bacteriophage gp41 helicase and gp61 primase assemble into a primosome to couple DNA unwinding with RNA primer synthesis for DNA replication. How the primosome is assembled and how the primer length is defined are unclear. Here we report a series of cryo-EM structures of T4 primosome assembly intermediates. We show that gp41 alone is an open spiral, and ssDNA binding triggers a large-scale scissor-like conformational change that drives the ring closure and activates the helicase. Helicase activation exposes a cryptic hydrophobic surface to recruit the gp61 primase. The primase binds the helicase in a bipartite mode in which the N-terminal Zn-binding domain and the C-terminal RNA polymerase domain each contain a helicase-interacting motif that bind to separate gp41 N-terminal hairpin dimers, leading to the assembly of one primase on the helicase hexamer. Our study reveals the T4 primosome assembly process and sheds light on the RNA primer synthesis mechanism.
Topics: Bacteriophage T4; DNA Primase; DNA Helicases; DNA Replication; DNA Primers; DNA, Viral
PubMed: 37474605
DOI: 10.1038/s41467-023-40106-2 -
Analytical Chemistry Dec 2023Partial DNA duplex formation greatly impacts the quality of DNA hybridization and has been extensively studied due to its significance in many biological processes....
Partial DNA duplex formation greatly impacts the quality of DNA hybridization and has been extensively studied due to its significance in many biological processes. However, traditional DNA sensing methods suffer from time-consuming amplification steps and hinder the acquisition of information about single-molecule behavior. In this work, we developed a plasmonic method to probe the hybridization process at a single base pair resolution and study the relationship between the complementarity of DNA analytes and DNA hybridization behaviors. We measured single-molecule hybridization events with Au NP-modified ssDNA probes in real time and found two hybridization adsorption events: stable and transient adsorption. The ratio of these two hybridization adsorption events was correlated with the length of the complementary sequences, distinguishing DNA analytes from different complementary sequences. By using dual incident angle excitation, we recognized different single-base complementary sequences. These results demonstrated that the plasmonic method can be applied to study partial DNA hybridization behavior and has the potential to be incorporated into the identification of similar DNA sequences, providing a sensitive and quantitative tool for DNA analysis.
Topics: Base Pairing; Nucleic Acid Hybridization; DNA; DNA, Single-Stranded; DNA Probes
PubMed: 38055795
DOI: 10.1021/acs.analchem.3c03316 -
Research (Washington, D.C.) 2024Photodynamic therapy (PDT) has emerged as a promising approach for squamous cell carcinoma treatment but hindered by tumor hypoxia, acquired resistance, phototoxicity,...
Photodynamic therapy (PDT) has emerged as a promising approach for squamous cell carcinoma treatment but hindered by tumor hypoxia, acquired resistance, phototoxicity, and so on. To address these issues, we developed a smart strategy utilizing activable photosensitizers delivered by an aptamer-functionalized DNA probe (ADP). The ADP incorporated an AS1411 aptamer for tumor targeting and a linear antisense oligonucleotide (ASO) for recognition of Survivin mRNA. In the absence of the target, PDT remained quenched, thereby avoiding phototoxicity during circulation and nonselective distribution. With the aid of the aptamer, ADP achieved selective targeting of tumors. Upon internalization, ADP targeted recognized Survivin mRNA, triggering PDT activation, and releasing ASO to down-regulate Survivin expression and reverse tumor resistance. Consequently, the activable photosensitizers exhibited an "AND" logic gate, combining tumor-targeting delivery and tumor-related gene activation, thus enhancing its specificity. Additionally, the incorporation of hemin into the ADP provided catalase activity, converting tumor-abundant HO into O, thereby ameliorating tumor hypoxia. The resulting functionalized G-quadruplex/hemin-DNA probe complex demonstrated targeted delivery and activation, minimized side effects, and enhanced PDT efficacy in both xenograft tumor-bearing mice and patient-derived xenograft models. This study offers a unique and promising platform for efficient and safe PDT, thus holding great potential for future clinical translation and improved cancer therapy.
PubMed: 38269029
DOI: 10.34133/research.0295 -
Analytical Chemistry Jan 2024The development of a simple, rapid, easy-to-operate, and ultrasensitive DNA walker-based sensing system is challenging but would be very intriguing for the enormous...
The development of a simple, rapid, easy-to-operate, and ultrasensitive DNA walker-based sensing system is challenging but would be very intriguing for the enormous applications in biological analysis and disease monitoring. Herein, a new self-propelled and self-enhanced DNA walking strategy was developed on the basis of a simple DNA polymerase-steered conversion from a typical alternate DNA assembly process. The sensing platform was fabricated easily by immobilizing only one hairpin probe (H1) and the sensing process was based on a simple one-step mixing with another hairpin-like DNA probe (H2) and DNA polymerase. The DNA polymerization could achieve target recycling and successive DNA walking steps. Interestingly, along with each DNA walking step, the new DNA walker sequence could be autonomously accumulated for a self-enhanced DNA walking effect. This provided a multilevel signal amplification ability for the ultrasensitive detection of the target with a low detection limit of 0.18 fM. Moreover, it could greatly reduce the reaction time with the sensing process finished within 1 h. The detection selectivity and the applicative potential in a complicated biological matrix were also demonstrated. Furthermore, the flexible control of sensing modes (self-enhanced DNA walking or the alternate DNA assembly) by using DNA polymerase or not offered a powerful means for sensing performance modulation. It thus opens a new avenue toward the development of a DNA walker-based sensing platform with both rapid and ultrasensitive features and might hold a huge potential for point-of-care diagnostic applications.
Topics: Biosensing Techniques; DNA; DNA Probes; DNA-Directed DNA Polymerase; Polymerization; Electrochemical Techniques; Limit of Detection
PubMed: 38158364
DOI: 10.1021/acs.analchem.3c04340 -
Analytical Chemistry Jul 2023We have developed a DNA sensor that can be finalized to detect a specific target on demand. The electrode surface was modified with 2,7-diamino-1,8-naphthyridine (DANP),...
We have developed a DNA sensor that can be finalized to detect a specific target on demand. The electrode surface was modified with 2,7-diamino-1,8-naphthyridine (DANP), a small molecule with nanomolar affinity for the cytosine bulge structure. The electrode was immersed in a solution of synthetic probe-DNA that had a cytosine bulge structure at one end and a complementary sequence to the target DNA at the other end. The strong binding between the cytosine bulge and DANP anchored the probe DNAs to the electrode surface, and the electrode became ready for target DNA sensing. The complementary sequence portion of the probe DNA can be changed as requested, allowing for the detection of a wide variety of targets. Electrochemical impedance spectroscopy (EIS) with the modified electrode detected target DNAs with a high sensitivity. The charge transfer resistance (Rct) extracted from EIS showed a logarithmic relationship with the concentration of target DNA. The limit of detection (LoD) was less than 0.01 μM. By this method, highly sensitive DNA sensors for various target sequences could be easily produced.
Topics: Dielectric Spectroscopy; Ligands; DNA; DNA Probes; Cytosine; Biosensing Techniques; Electrodes; Electrochemical Techniques
PubMed: 37341999
DOI: 10.1021/acs.analchem.3c01126 -
ACS Applied Bio Materials Jan 2024Cancers remain the leading cause of mortality worldwide. It is crucial to detect cancer at an early stage for improving survival rates. Biomarkers have precise...
Cancers remain the leading cause of mortality worldwide. It is crucial to detect cancer at an early stage for improving survival rates. Biomarkers have precise implications for cancer progression. Here, we built a straightforward DNA probe system that could be activated by near-infrared light to detect dual miRNAs with a high specificity. This probe is built on the basis of upconversion nanoparticles, which could emit ultraviolet light and activate DNA probes adsorbed on the outer layer. The DNA probe system is remotely controlled through manipulation of the near-infrared (NIR) light, enabling simultaneous detection of dual miRNAs. The DNA nanosystem could be effectively endocytosed by cancer cells and reflect expression levels of dual miRNAs. Overall, this study demonstrates a promising remote-controlled DNA nanoplatform for the simultaneous detection of dual miRNAs, which has tremendous potential for precise cancer diagnostics and therapies.
Topics: Humans; MicroRNAs; Ultraviolet Rays; DNA; DNA Probes; Nanoparticles; Neoplasms
PubMed: 38151236
DOI: 10.1021/acsabm.3c01079 -
Scientific Reports Jan 2024mRNA measurement is dominated by RT-PCR, which requires expensive laboratory equipment and personnel with advanced degrees. Loop-mediated isothermal amplification (LAMP)...
mRNA measurement is dominated by RT-PCR, which requires expensive laboratory equipment and personnel with advanced degrees. Loop-mediated isothermal amplification (LAMP) is a versatile technique for detecting target DNA and RNA. The sensitivity of LAMP in early reports has been below that of the standard RT-PCR tests. Here, we report the use of a fluorescence-based RT-LAMP protocol to measure CDX2 expression patterns, which match extremely well to the standards of sophisticated RT-PCR techniques (r = 0.99, p < 0.001). The assay works on diverse sample types such as cDNA, mRNA, and direct tissue sample testing in 25 min compared to more than 3 h for RT-PCR. We have developed a new protocol for designing RT-LAMP primers that reduce false positives due to self-amplification and improve quantification. A simple device with a 3D-printed box enables the measurement of mRNA expression at home, outdoors, and point-of-care setting.
Topics: RNA, Messenger; RNA; Biological Assay; DNA Primers; DNA, Complementary
PubMed: 38200031
DOI: 10.1038/s41598-023-49651-8