DNA Tiles Research Articles

Programmed DNA tile response system enabled accurate detection of ATP for clinical diagnosis

Adenosine triphosphate (ATP), as a crucial signaling molecule, is of high potential diagnosis value for multiple diseases. However, current technologies for ATP detection struggle to satisfy the harsh conditions required for biospecimen tests, existing several limitations for clinical practical application. In this study, the DNA tile response system was designed and constructed by integrating nucleic acid aptamer, hybrid chain reaction (HCR), and DNA tile self-assembly technology to achieve accurate detection of ATP in body fluid specimens. The DNA tile response system, possessing high-order structure characteristic, could remove HCR false-positive products and conquer the high noise background to enhance the detection sensitivity. Under the optimal reaction parameters, this system achieved the low limit of detection (LOD) of 189.23 nM over a wide linear range and splendid detection specificity faced with multiple interference terms. Furthermore, this system, possessing simple storage conditions, long-term detection stability, and superior anti-interference performance, was sufficient to satisfy the detection requirements for complicated biological specimens in multi-scenarios. Ultimately, the DNA tile response system showed excellent diagnostic efficiency that could easily distinguish urinary tract infection patients and healthy controls through ATP quantification, as well as hypertension patients and healthy ones. This system provides a high-potential rapid detection tool for early diagnosis and dynamic monitoring of urinary tract infections and hypertension, holding enormous potential for clinical translation due to the superiorities of simple operation, general applicability, and low instrument requirements.

Developmental assembly of multi-component polymer systems through interconnected synthetic gene networks in vitro

Living cells regulate the dynamics of developmental events through interconnected signaling systems that activate and deactivate inert precursors. This suggests that similarly, synthetic biomaterials could be designed to develop over time by using chemical reaction networks to regulate the availability of assembling components. Here we demonstrate how the sequential activation or deactivation of distinct DNA building blocks can be modularly coordinated to form distinct populations of self-assembling polymers using a transcriptional signaling cascade of synthetic genes. Our building blocks are DNA tiles that polymerize into nanotubes, and whose assembly can be controlled by RNA molecules produced by synthetic genes that target the tile interaction domains. To achieve different RNA production rates, we use a strategy based on promoter “nicking” and strand displacement. By changing the way the genes are cascaded and the RNA levels, we demonstrate that we can obtain spatially and temporally different outcomes in nanotube assembly, including random DNA polymers, block polymers, and as well as distinct autonomous formation and dissolution of distinct polymer populations. Our work demonstrates a way to construct autonomous supramolecular materials whose properties depend on the timing of molecular instructions for self-assembly, and can be immediately extended to a variety of other nucleic acid circuits and assemblies.

Binding-induced activation of CRISPR-Cas12a using PAM-engineered toehold switch and DNA tile architectures for enhanced detection of thrombin

This study presents a novel biosensor that integrates CRISPR-Cas12a technology with DNA tile-based architectures to enhance thrombin detection. Utilizing PAM-engineered toehold switches, the biosensor dynamically activates the CRISPR-Cas12a system upon thrombin recognition, significantly improving both sensitivity and specificity. The label-free electrochemical detection method offers rapid, cost-effective analysis, making it ideal for real-time clinical applications. The biosensor achieved an impressive detection limit of 103 aM, surpassing traditional methods and setting a new standard for thrombin assays. The biosensor's robust performance in complex biological samples, such as diluted human serum, demonstrates its reliability for clinical diagnostics. This is particularly important for the early diagnosis and management of thrombotic and coagulation-related disorders. By combining advanced genetic engineering with traditional electrochemical techniques, the biosensor not only optimizes detection but also expands its potential applications across various biomedical fields. Overall, this biosensor provides a powerful and innovative tool for thrombin detection, offering enhanced diagnostic accuracy, faster patient management, and potential cost savings in healthcare. Its superior performance and adaptability highlight its promise for broader adoption in clinical diagnostics and other biosensing applications.

A biomimetic branching signal-passing tile assembly model with dynamic growth and disassembly.

Natural biological branching processes can form tree-like structures at all scales and, moreover, can perform various functions to achieve specific goals; these include receiving stimuli, performing two-way communication along their branches, and dynamically reforming (extending or retracting branches). They underlie many biological systems with considerable diversity, frequency, and geometric complexity; these include networks of neurons, organ tissue, mycorrhizal fungal networks, plant growth, foraging networks, etc. This paper presents a biomimetic DNA tile assembly model (Y-STAM) to implement dynamic branching processes. The Y-STAM is a relatively compact mathematical model providing a design space where complex, biomimetic branch-like growth and behaviour can emerge from the appropriate parametrization of the model. We also introduce a class of augmented models (Y-STAM+) that provide time- and space-dependent modulations of tile glue strengths, which enable further diverse behaviours that are not possible in the Y-STAM; these additional behaviours include refinement of network assemblies, obstacle avoidance, and programmable growth patterns. We perform and discuss extensive simulations of the Y-STAM and the Y-STAM+. We envision that these models could be applied at the mesoscale and the molecular scale to dynamically assemble branching DNA nanostructures and offer insights into complex biological self-assembly processes.

Diverse Chiral Nanotubes Assembled from Identical DNA Strands.

DNA nanotubes with controllable geometries hold a wide range of interdisciplinary applications. When preparing DNA nanotubes of varying widths or distinct chirality, existing methods require repeatedly designing and synthesizing specific DNA sequences, which can be costly and laborious. Here, we proposed an intercalator-assisted DNA tile assembly method which enables the production of DNA nanotubes of diverse widths and chirality using identical DNA strands. Through adjusting the concentration of intercalators during assembly, the twisting direction and extent of DNA tiles could be modulated, leading to the formation of DNA nanotubes featuring controllable widths and chirality. Moreover, through introducing additional intercalators and secondary annealing, right-handed nanotubes could be reconfigured into distinct left-handed nanotubes. We expect that this method could be universally applied to modulating the self-assembly pathways of various DNA tiles and other chiral materials, advancing the landscape of DNA tile assembly.

Cascadable-Controllable Self-Assembly DNA Tiles for Large-Scale DNA Logic Circuits.

In the last few decades, DNA-based self-assembly tiles has become a hot field in research due to its special applications and advantages. The regularity and strong design methods comprise other DNA-based digital circuit design methods. In addition to the obvious advantages of this method, there are challenges in performing computations based on self-assembly tiles, which have hindered the development and construction of large computing circuits with this method. The first challenge is the creation of crystals from DNA molecules in the output, which has led to the impossibility of cascading. The second challenge of this method is the uncontrollability of the reactions of the tiles, which increases the percentage of computing errors. In this article, these two challenges have been solved by changing the structure of leading tiles so that without the activator strand, tiles remain inactive and cannot be connected to other tiles. Also, when the tiles are activated, single-strand DNA will be released after connecting to other tiles, which will be used as the output of the circuit. This output gives the possibility of cascading to self-assembly designed circuits. The method introduced in this article can be a beginning for the re-development of DNA-based circuit design with the self-assembly tile method.

Three-Point-Star Deoxyribonucleic Acid Tiles with the Core Arm Length at Three Half-Turns for Two-Dimensional Archimedean Tilings and Beyond.

2D Archimedean tiling and complex tessellation patterns assembled from soft materials including modular DNA tiles have attracted great interest because of their specific structures and potential applications in nanofabrication, nanoelectronics, nanophotonics, biomedical sensing, drug delivery, therapeutics, etc. Traditional three- and four-point-star DNA tiles with the core arm length at two half-turns (specified as three- and four-point-star-E previously and abbreviated as 3PSE and 4PSE tiles here) have been applied to assemble intricate tessellations through tuning the size of inserted nT (n = 1-7, T is thymine) loops on helper strands at the tile center. Following our recent findings using a new type of four-point-star tiles with the core arm length at three half-turns (specified as four-point-star-O previously and abbreviated as 4PSO tiles here) to assemble DNA tubes and flat 2D arrays, we report here the cross-hybridization weaving architectures at the tile center to construct three new 3PSO tiles with circular DNA oligonucleotides of 96-nt (nucleotides) serving as the scaffolds, further the monotonous and combinatory E- and O-tilings on one type of 3PSO tiles to create 2D Archimedean tiling patterns (6.6.6) and (4.8.8), and finally, the combination of 3PSO with 4PSO as well as 2PSO tiles to tile into complex tessellation patterns. The easy realization of regular and intricate DNA tessellations with 2-4PSO tiles not only richens the fundamental DNA modules and complex DNA nanostructures in types but also broadens the potential application scopes of DNA nanostructures in nanofabrication, DNA computing, biomedicine, etc.

Synthetic DNA-based Swimmers Driven by Enzyme Catalysis.

Here, we report DNA-based synthetic nanostructures decorated with enzymes (hereafter referred to as DNA-enzyme swimmers) that self-propel by converting the enzymatic substrate to the product in solution. The DNA-enzyme swimmers are obtained from tubular DNA structures that self-assemble spontaneously by the hybridization of DNA tiles. We functionalize these DNA structures with two different enzymes, urease and catalase, and show that they exhibit concentration-dependent movement and enhanced diffusion upon addition of the enzymatic substrate (i.e., urea and H2O2). To demonstrate the programmability of such DNA-based swimmers, we also engineer DNA strands that displace the enzyme from the DNA scaffold, thus acting as molecular "brakes" on the DNA swimmers. These results serve as a first proof of principle for the development of synthetic DNA-based enzyme-powered swimmers that can self-propel in fluids.

Guided Self-Assembly of DNA Tiles into One- and Two-Dimensional Patterns Using Strand-Displacement and Optochemical Pathways

Guided Self-Assembly of DNA Tiles into One- and Two-Dimensional Patterns Using Strand-Displacement and Optochemical Pathways

Y-Shaped Backbone-Rigidified DNA Tiles for the Construction of Supersized Nondeformable Tetrahedrons for Precise Cancer Therapies.

While engineered DNA nanoframeworks have been extensively exploited for delivery of diagnostic and therapeutic regents, DNA tiling-based DNA frameworks amenable to applications in living systems lag much behind. In this contribution, by developing a Y-shaped backbone-based DNA tiling technique, we assemble Y-shaped backbone-rigidified supersized DNA tetrahedrons (RDT) with 100% efficiency for precisely targeted tumor therapy. RDT displays unparalleled rigidness and unmatched resistance to nuclease degradation so that it almost does not deform under the force exerted by the atomic force microscopy tip, and the residual amount is not less than 90% upon incubating in biological media for 24 h, displaying at least 11.6 times enhanced degradation resistance. Without any targeting ligand, RDT enters the cancer cell in a targeted manner, and internalization specificity is up to 15.8. Moreover, 77% of RDT objects remain intact within living cells for 14 h. The drug loading content of RDT is improved by 4-8 times, and RDT almost 100% eliminates the unintended drug leakage in a stimulated physiological medium. Once systemically administrated into HeLa tumor-bearing mouse models, doxorubicin-loaded RDTs preferentially accumulate in tumor sites and efficiently suppress tumor growth without detectable off-target toxicity. The Y-DNA tiling technique offers invaluable insights into the development of structural DNA nanotechnology for precise medicine.

Pattern recognition in the nucleation kinetics of non-equilibrium self-assembly

Inspired by biology’s most sophisticated computer, the brain, neural networks constitute a profound reformulation of computational principles1–3. Analogous high-dimensional, highly interconnected computational architectures also arise within information-processing molecular systems inside living cells, such as signal transduction cascades and genetic regulatory networks4–7. Might collective modes analogous to neural computation be found more broadly in other physical and chemical processes, even those that ostensibly play non-information-processing roles? Here we examine nucleation during self-assembly of multicomponent structures, showing that high-dimensional patterns of concentrations can be discriminated and classified in a manner similar to neural network computation. Specifically, we design a set of 917 DNA tiles that can self-assemble in three alternative ways such that competitive nucleation depends sensitively on the extent of colocalization of high-concentration tiles within the three structures. The system was trained in silico to classify a set of 18 grayscale 30 × 30 pixel images into three categories. Experimentally, fluorescence and atomic force microscopy measurements during and after a 150 hour anneal established that all trained images were correctly classified, whereas a test set of image variations probed the robustness of the results. Although slow compared to previous biochemical neural networks, our approach is compact, robust and scalable. Our findings suggest that ubiquitous physical phenomena, such as nucleation, may hold powerful information-processing capabilities when they occur within high-dimensional multicomponent systems.

Protonation-mediated DNA tile self-assembly with nuclease resistance characteristic for signal-amplified detection of microRNAs

DNA nanotechnology, developing rapidly in recent years, has unprecedented superiorities in biological application-oriented research including high programmability, convenient functionalization, reconfigurable structure, and intrinsic biocompatibility. However, the susceptibility to nucleases in the physiological environment has been an obstacle to applying DNA nanostructures in biological science research. In this study, a new DNA self-assembly strategy, mediated by double-protonated small molecules instead of classical metal ions, is developed to enhance the nuclease resistance of DNA nanostructures while retaining their integrality and functionality, and the relative application has been launched in the detection of microRNAs (miRNAs). Faced with low-abundance miRNAs, we integrate hybrid chain reaction (HCR) with DNA self-assembly in the presence of double-protonated small molecules to construct a chemiluminescence detection platform with nuclease resistance, which utilizes the significant difference of molecular weight between DNA arrays and false-positive products to effectively separate of reaction products and remove the detection background. This strategy attaches importance to the nucleic acid stability during the assay process via improving nuclease resistance while rendering the detection results for miRNAs more authentic and reliable, opening our eyes to more possibilities for the multiple applications of customized DNA nanostructures in biology, including bioassay, bioimaging, drug delivery, and cell modulation.

DNA T-shaped crossover tiles for 2D tessellation and nanoring reconfiguration

DNA tiles serve as the fundamental building blocks for DNA self-assembled nanostructures such as DNA arrays, origami, and designer crystals. Introducing additional binding arms to DNA crossover tiles holds the promise of unlocking diverse nano-assemblies and potential applications. Here, we present one-, two-, and three-layer T-shaped crossover tiles, by integrating T junction with antiparallel crossover tiles. These tiles carry over the orthogonal binding directions from T junction and retain the rigidity from antiparallel crossover tiles, enabling the assembly of various 2D tessellations. To demonstrate the versatility of the design rules, we create 2-state reconfigurable nanorings from both single-stranded tiles and single-unit assemblies. Moreover, four sets of 4-state reconfiguration systems are constructed, showing effective transformations between ladders and/or rings with pore sizes spanning ~20 nm to ~168 nm. These DNA tiles enrich the design tools in nucleic acid nanotechnology, offering exciting opportunities for the creation of artificial dynamic DNA nanopores.

Design of DNA-Based Artificial Transmembrane Channels for Biosensing and Biomedical Applications

Biomolecular channels on the cell membrane are essential for transporting substances across the membrane to maintain cell physiological activity. Artificial transmembrane channels used to mimic biological membrane channels can regulate intra/extracellular ionic and molecular homeostasis, and they elucidate cellular structures and functionalities. Due to their program design, facile preparation, and high biocompatibility, DNA nanostructures have been widely used as scaffolds for the design of artificial transmembrane channels and exploited for ionic and molecular transport and biomedical applications. DNA-based artificial channels can be designed from two structural modules: DNA nanotubes/nanopores as transport modules for mass transportation and hydrophobic segments as anchor modules for membrane immobilization. In this review, various lipophilic modification strategies for the design of DNA channels and membrane insertion are outlined. Several types of DNA transmembrane channels are systematically summarized, including DNA wireframe channels, DNA helix bundle channels, DNA tile channels, DNA origami channels, and so on. We then discuss efforts to exploit them in biosensor and biomedical applications. For example, ligand-gated and environmental stimuli-responsive artificial transmembrane channels have been designed for transmembrane signal transduction. DNA-based artificial channels have been developed for cell mimicry and the regulation of cell behaviors. Finally, we provide some perspectives on the challenges and future developments of artificial transmembrane channel research in biomimetic science and biomedical applications.

AMP Aptamer Programs DNA Tile Cohesion without Canonical Base Pairing.

Tile-based DNA self-assembly provides a versatile approach for the construction of a wide range of nanostructures for various applications such as nanomedicine and advanced materials. The inter-tile interactions are primarily programmed by base pairing, particularly Watson-Crick base pairing. To further expand the tool box for DNA nanotechnology, herein, we have designed DNA tiles that contain both ligands and aptamers. Upon ligand-aptamer binding, tiles associate into geometrically well-defined nanostructures. This strategy has been demonstrated by the assembly of a series of DNA nanostructures, which have been thoroughly characterized by gel electrophoresis and atomic force microscopy. This new inter-tile cohesion could bring new potentials to DNA self-assembly in the future. For example, the addition of free ligand could modulate the nanostructure formation. In the case of biological ligands, DNA self-assembly could be related to the presence of certain ligands.

DNA Tile and Invading Stacking Primer-Assisted CRISPR-Cas12a Multiple Amplification System for Entropy-Driven Electrochemical Detection of MicroRNA with Tunable Sensitivity.

Conventional electrochemical detection of microRNA (miRNA) encounters issues of poor sensitivity and fixed dynamic range. Here, we report a DNA tile and invading stacking primer-assisted CRISPR-Cas12a multiple amplification strategy to construct an entropy-controlled electrochemical biosensor for the detection of miRNA with tunable sensitivity and dynamic range. To amplify the signal, a cascade amplification of the CRISPR-Cas12a system along with invading stacking primer signal amplification (ISPSA) was designed to detect trace amounts of miRNA-31 (miR-31). The target miR-31 could activate ISPSA and produce numerous DNAs, triggering the cleavage of the single-stranded linker probe (LP) that connects a methylene blue-labeled DNA tile with a DNA tetrahedron to form a Y-shaped DNA scaffold on the electrode. Based on the decrease of current, miR-31 can be accurately and efficiently detected. Impressively, by changing the loop length of the LP, it is possible to finely tune the entropic contribution while keeping the enthalpic contribution constant. This strategy has shown a tunable limit of detection for miRNA from 0.31 fM to 0.56 pM, as well as a dynamic range from ∼2200-fold to ∼270,000-fold. Moreover, it demonstrated satisfactory results in identifying cancer cells with a high expression of miR-31. Our strategy broadens the application of conventional electrochemical biosensing and provides a tunable strategy for detecting miRNAs at varying concentrations.

Recent Advances in DNA Nanotechnology-Enabled Biosensors for Virus Detection.

Virus-related infectious diseases are serious threats to humans, which makes virus detection of great importance. Traditional virus-detection methods usually suffer from low sensitivity and specificity, are time-consuming, have a high cost, etc. Recently, DNA biosensors based on DNA nanotechnology have shown great potential in virus detection. DNA nanotechnology, specifically DNA tiles and DNA aptamers, has achieved atomic precision in nanostructure construction. Exploiting the programmable nature of DNA nanostructures, researchers have developed DNA nanobiosensors that outperform traditional virus-detection methods. This paper reviews the history of DNA tiles and DNA aptamers, and it briefly describes the Baltimore classification of virology. Moreover, the advance of virus detection by using DNA nanobiosensors is discussed in detail and compared with traditional virus-detection methods. Finally, challenges faced by DNA nanobiosensors in virus detection are summarized, and a perspective on the future development of DNA nanobiosensors in virus detection is also provided.

Molecular Lithography on Silicon Wafers Guided by Porous, Extended Arrays of Small DNA Tiles.

Tile-based DNA self-assembly is a cost-effective fabrication method for large-scale nanopatterns. Herein, we report a protocol to directly assemble DNA 2D arrays on silicon wafers and then use the DNA nanostructures as molds to fabricate the corresponding nanostructures on the silicon wafers by hydrogen fluoride (HF) etching. Similar HF etching has been used with robust large DNA origami structures as templates. This work demonstrates that DNA nanostructures assembled from small tiles are sufficiently stable for this process. The resulting feature size (∼8.6 nm) approaches the sizes of e-beam lithography. While the reported method is parallel and inexpensive, e-beam lithography is a serial method and is expensive. We expect that this method will be very useful for preparing fine nanopatterns in large areas.

Integrative modeling of lncRNA-chromatin interaction maps reveals diverse mechanisms of nuclear retention

BackgroundMany long non-coding RNAs, known to be involved in transcriptional regulation, are enriched in the nucleus and interact with chromatin. However, their mechanisms of chromatin interaction and the served cellular functions are poorly understood. We sought to characterize the mechanisms of lncRNA nuclear retention by systematically mapping the sequence and chromatin features that distinguish lncRNA-interacting genomic segments.ResultsWe found DNA 5-mer frequencies to be predictive of chromatin interactions for all lncRNAs, suggesting sequence-specificity as a global theme in the interactome. Sequence features representing protein-DNA and protein-RNA binding motifs revealed potential mechanisms for specific lncRNAs. Complementary to these global themes, transcription-related features and DNA-RNA triplex formation potential were noted to be highly predictive for two mutually exclusive sets of lncRNAs. DNA methylation was also noted to be a significant predictor, but only when combined with other epigenomic features.ConclusionsTaken together, our statistical findings suggest that a group of lncRNAs interacts with transcriptionally inactive chromatin through triplex formation, whereas another group interacts with transcriptionally active regions and is involved in DNA Damage Response (DDR) through formation of R-loops. Curiously, we observed a strong pattern of enrichment of 5-mers in four potentially interacting entities: lncRNA-bound DNA tiles, lncRNAs, miRNA seed sequences, and repeat elements. This finding points to a broad sequence-based network of interactions that may underlie regulation of fundamental cellular functions. Overall, this study reveals diverse sequence and chromatin features related to lncRNA-chromatin interactions, suggesting potential mechanisms of nuclear retention and regulatory function.

Building Large DNA Bundles via Controlled Hierarchical Assembly of DNA Tubes.

Structural DNA nanotechnology is capable of fabricating designer nanoscale artificial architectures. Developing simple and yet versatile assembly methods to construct large DNA structures of defined spatial features and dynamic capabilities has remained challenging. Herein, we designed a molecular assembly system where DNA tiles can assemble into tubes and then into large one-dimensional DNA bundles following a hierarchical pathway. A cohesive link was incorporated into the tile to induce intertube binding for the formation of DNA bundles. DNA bundles with length of dozens of micrometers and width of hundreds of nanometers were produced, whose assembly was revealed to be collectively determined by cationic strength and linker designs (binding strength, spacer length, linker position, etc.). Furthermore, multicomponent DNA bundles with programmable spatial features and compositions were realized by using various distinct tile designs. Lastly, we implemented dynamic capability into large DNA bundles to realize reversible reconfigurations among tile, tube, and bundles following specific molecular stimulations. We envision this assembly strategy can enrich the toolbox of DNA nanotechnology for rational design of large-size DNA materials of defined features and properties that may be applied to a variety of fields in materials science, synthetic biology, biomedical science, and beyond.