Stepwise Assembly of DNA Nanostructures in a Surface-Based Method.

Hierarchical assembly of DNA nanostructures has already led to superstructures of ever-increasing level of complexity. Processing control in building nanostructures hierarchically is desirable but remains underexplored. Here, we present the stepwise assembly of DNA origami nanostructures by a surface-based method.

Expanding DNA Origami Design Freedom with De Novo Synthesized Scaffolds.

The construction of DNA origami nanostructures is heavily dependent on the folding of the scaffold strand, which is typically a single-stranded DNA genome extracted from a bacteriophage (M13). Custom scaffolds can be prepared in a number of methods, but they are not widely accessible to a broad user base in the DNA nanotechnology community. Here, we explored new design and construction possibilities with custom scaffolds prepared in our cost- and time-efficient production pipeline.

DNA Self-Assembly Optimization by Betaine and Its Analogs.

The self-assembly yield of DNA nanostructures can be exponentially lower with increasing structural complexity. Few optimizing strategies are available in the DNA nanotechnology field for the assembly yield improvement. Here, betaine and its analogs are applied as supplementary ingredients in DNA self-assembly.

Synthetic molecular switches driven by DNA-modifying enzymes.

Taking inspiration from natural systems, in which molecular switches are ubiquitous in the biochemistry regulatory network, we aim to design and construct synthetic molecular switches driven by DNA-modifying enzymes, such as DNA polymerase and nicking endonuclease. The enzymatic treatments on our synthetic DNA constructs controllably switch ON or OFF the sticky end cohesion and in turn cascade to the structural association or disassociation. Here we showcase the concept in multiple DNA nanostructure systems with robust assembly/disassembly performance.

Enzymatic Assembly of DNA Nanostructures and Fragments with Sequence Overlaps.

Homologous recombination, an evolutionarily conserved DNA double-strand break repair pathway to protect genome stability, has long been exploited for the and assembly of multiple DNA duplex fragments in molecular cloning. Whether such methods can also be applied in the self-assembly of DNA nanostructures remains underexplored. Here, we report an enzymatic approach for the self-assembly of high-order DNA constructs with overlapping segments.

Controllable dynamics of complex DNA nanostructures.

In the past four decades, a variety of self-assembly design frameworks have led to the construction of versatile DNA nanostructures with increasing complexity and controllability. The controllable dynamics of DNA nanostructures has garnered much interest and emerged as a powerful tool for conducting sophisticated tasks at the molecular level. In this minireview, we summarized the controllable reconfigurations of complex DNA nanostructures induced by nucleic acid strands, environmental stimuli and enzymatic treatments.

Mesojunction-Based Design Paradigm of Structural DNA Nanotechnology.

Mesojunctions were introduced as a basic type of crossover configuration in the early development of structural DNA nanotechnology. However, the investigations of self-assembly from multiple mesojunction complexes have been overlooked in comparison to their counterparts based on regular junctions. In this work, we designed standardized component strands for the construction of complex mesojunction lattices.

Design of Orthogonal DNA Sticky-End Cohesion Based on Configuration-Specific Molecular Recognition.

In sticky-end cohesion, sequence complementarity is the key to design specific recognition among many DNA nanostructure units. The binding orthogonality usually arises from sticky-end pairs of different sequences. Instead of creating orthogonal species of sticky-end bonds based on sequence complementarity, we restricted the sticky-end sequence diversity down to the fixed C-G pair and explored orthogonal recognition of the synthetic DNA constructs based solely on the configurational match.

Reconfiguration of DNA nanostructures induced by enzymatic ligation treatment.

Enzymatic ligation is a popular method in DNA nanotechnology for structural enforcement. When employed as stability switch for chosen components, ligation can be applied to induce DNA nanostructure reconfiguration. In this study, we investigate the reinforcement effect of ligation on addressable DNA nanostructures assembled entirely from short synthetic strands as the basis of structural reconfiguration.

Controllable assembly of synthetic constructs with programmable ternary DNA interaction.

Compared with the dual binding components in a binary interaction, the third component of a ternary interaction often serves as modulator or regulator in biochemical processes. Here, we presented a programmable ternary interaction strategy based on the natural DNA triplex structure. With the DNA triplex-based ternary interaction, we have successfully demonstrated controllable hierarchical assemblies from nanometer scale synthetic DNA nanostructure units to micrometer scale live bacteria.

Rational Design of Allosteric Nanodevices Based on DNA Triple Helix.

Inspired by allosteric regulation of natural molecules, we present a rational design scheme to build synthetic nucleic acid allosteric nanodevices. The clearly specified conformational states of switches obtained from systematic screening and analyses make the ON-OFF transition clear-cut and quantification ready. Under the rational design scheme, we have developed a series of DNA switches with triplex-forming oligos as allosteric modulators and implemented designated allosteric transitions, allosteric coregulation, and reaction pathway control.

Reconfigurable Two-Dimensional DNA Lattices: Static and Dynamic Angle Control.

Branched DNA motifs serve as the basic construction elements for all synthetic DNA nanostructures. However, precise control of branching orientation remains a key challenge to further heighten the overall structural order. In this study, we use two strategies to control the branching orientation.

Monochromatic Fluorescent Barcodes Hierarchically Assembled from Modular DNA Origami Nanorods.

With the rapid advancement of fluorescence microscopy, there is a growing interest in the multiplexed detection and identification of various bioanalytes (e.g., nucleic acids and proteins) for efficient sample processing and analysis.

DNA dynamics and computation based on toehold-free strand displacement.

We present a simple and effective scheme of a dynamic switch for DNA nanostructures. Under such a framework of toehold-free strand displacement, blocking strands at an excess amount are applied to displace the complementation of specific segments of paired duplexes. The functional mechanism of the scheme is illustrated by modelling the base pairing kinetics of competing strands on a target strand.

Hybrid Wireframe DNA Nanostructures with Scaffolded and Scaffold-Free Modules.

The scaffolded DNA origami approach and the scaffold-free LEGO approach both demonstrate an extraordinary self-assembly capability for constructing all kinds of complex DNA nanostructures. Combining the construction elements of the two approaches, we introduce a hybrid framework to build wireframe structures in this study. A collection of two-dimensional (2D) and three-dimensional (3D) wireframe structures are presented to showcase the simple and versatile design.

Allostery of DNA nanostructures controlled by enzymatic modifications.

Allostery is comprehensively studied for natural macromolecules, such as proteins and nucleic acids. Here, we present controllable allostery of synthetic DNA nanostructure-enzyme systems. Rational designs of the synthetic allosteric systems are based on an in-depth understanding of allosteric sites with several types of strand placements, whose varying stacking strengths determine the local conformation and ultimately lead to a gradient level of allosteric transition.

Addressable DNA nanotubes with repetitive components.

Extended DNA nanostructures have already been constructed in a repetitive arrangement from millions of building blocks, many more than currently feasible with even the gold standard of addressable self-assembled structures. In order to construct addressable DNA nanostructures with more building blocks, it is desirable to arrange the addressable components repetitively. Accordingly, the overall size of the structure can be multiplied by the level of repetition in the addressable strands.

DNA nanostructures from double-C-shaped motifs with controllable twist and curvature.

We demonstrate twist and curvature engineering in DNA nanostructures from the scaffold-free approach. The DNA 'LEGO' bricks adopted in this study are double-C-shaped motifs, and extended nanostructures are constructed to visualize the structural details of twist or curvature. By systematically deleting and inserting base pairs at certain domains of the component motifs, we are able to study various levels of the twist and curvature of the resulting nanostructures comprehensively.

Self-Assembly of Wireframe DNA Nanostructures from Junction Motifs.

Wireframe frameworks have been investigated for the construction of complex nanostructures from a scaffolded DNA origami approach; however, a similar framework is yet to be fully explored in a scaffold-free "LEGO" approach. Herein, we describe a general design scheme to construct wireframe DNA nanostructures entirely from short synthetic strands. A typical edge of the resulting structures in this study is composed of two parallel duplexes with crossovers on both ends, and three, four, or five edges radiate out from a certain vertex.

Complex wireframe DNA nanostructures from simple building blocks.

DNA nanostructures with increasing complexity have showcased the power of programmable self-assembly from DNA strands. At the nascent stage of the field, a variety of small branched objects consisting of a few DNA strands were created. Since then, a quantum leap of complexity has been achieved by a scaffolded 'origami' approach and a scaffold-free approach using single-stranded tiles/bricks-creating fully addressable two-dimensional and three-dimensional DNA nanostructures designed on densely packed lattices.

Hierarchical Assembly of DNA Nanostructures Based on Four-Way Toehold-Mediated Strand Displacement.

Because of its attractive cost and yield, hierarchical assembly, in which constituent structures of lower hierarchy share a majority of components, is an appealing approach to scale up DNA self-assembly. A few strategies have already been investigated to combine preformed DNA nanostructures. In this study, we present a new hierarchical assembly method based on four-way toehold-mediated strand displacement to facilitate the combination of preformed DNA structural units.

Sub-100-nm metafluorophores with digitally tunable optical properties self-assembled from DNA.

Fluorescence microscopy allows specific target detection down to the level of single molecules and has become an enabling tool in biological research. To transduce the biological information to an imageable signal, we have developed a variety of fluorescent probes, such as organic dyes or fluorescent proteins with different colors. Despite their success, a limitation on constructing small fluorescent probes is the lack of a general framework to achieve precise and programmable control of critical optical properties, such as color and brightness.

Versatile DNA Origami Nanostructures in Simplified and Modular Designing Framework.

We introduce a simplified and modular architecture for design and construction of complex origami nanostructures. A series of basic two-dimensional and three-dimensional structures are presented. As the resulting structures can be virtually divided into blocks, modular remodeling such as translocation, contraction/extension, and bending is carried out.

DNA nanostructures constructed with multi-stranded motifs.

Earlier studies in DNA self-assembly have foretold the feasibility of building addressable nanostructures with multi-stranded motifs, which is fully validated in this study. In realizing this feasibility in DNA nanotechnology, a diversified set of motifs of modified domain lengths is extended from a classic type. The length of sticky ends can be adjusted to form different dihedral angles between the matching motifs, which corresponds to different connecting patterns.

Self-assembly of fully addressable DNA nanostructures from double crossover tiles.

DNA origami and single-stranded tile (SST) are two proven approaches to self-assemble finite-size complex DNA nanostructures. The construction elements appeared in structures from these two methods can also be found in multi-stranded DNA tiles such as double crossover tiles. Here we report the design and observation of four types of finite-size lattices with four different double crossover tiles, respectively, which, we believe, in terms of both complexity and robustness, will be rival to DNA origami and SST structures.