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This manuscript introduces geometry as a means to program the tile-based DNA self-assembly in two and three dimensions. This strategy complements the sequence-focused programmable assembly. DNA crystal assembly critically relies on intermotif, sticky-end cohesion, which requires complementarity not only in sequence but also in geometry. For DNA motifs to assemble into crystals, they must be associated with each other in the proper geometry and orientation to ensure that geometric hindrance does not prevent sticky ends from associating. For DNA motifs with exactly the same pair of sticky-end sequences, by adjusting the length (thus, helical twisting phase) of the motif branches, it is possible to program the assembly of these distinct motifs to either mix with one another, to self-sort and consequently separate from one another, or to be alternatingly arranged. We demonstrate the ability to program homogeneous crystals, DNA "alloy" crystals, and definable grain boundaries through self-assembly. We believe that the integration of this strategy and conventional sequence-focused assembly strategy could further expand the programming versatility of DNA self-assembly.

The overwinding and underwinding of duplex segments between junctions have been used in designing both left-handed and right-handed DNA origami nanostructures. For a variety of DNA tubes obtained from self-assembled tiles, only a theoretical approach of the intrinsic curvature of the DNA tile (specified as the intrinsic tile curvature) has been previously used to explain their formation. Details regarding the quantitative and structural descriptions of the tile curvature and its evolution in DNA tubes by the coupling of the twist of the inter-tile arm (specified as the arm twist) have never been addressed. In this work, we designed three types of tile cores built around a circular 128 nucleotide scaffold by using longitudinal weaving (LW), bridged longitudinal weaving (bLW) and transverse weaving (TW). Joining the tiles with inter-tile arms having the length of an odd number of DNA half-turns (termed O-tiling) almost resulted into planar 2D lattices, whereas joining the tiles with the arms having the length of an even number of DNA half-turns (termed E-tiling) nearly generated tubes. Streptavidin bound to biotin was used as a labeling technique to characterize the inside and outside surfaces of the E-tiling tubes and thereby the conformations of their component tiles with addressable concave and convex curvatures. When the arms have the normal winding at the relaxed B-form of DNA, the intrinsic tile curvature deter-mines the chirality of the E-tiling tubes. By regulating the arm length and the sticky end length of the bLW-Ep/q (E-tiling of the bLW cores with the arm length of p-bp and the sticky end length of q-nt) assemblies, the arm can be overwound, resulting in a left-handed twist, and can also be underwound, resulting in a right-handed twist. Chiral bLW-Ep/q tubes with either a right-handed curvature or a left-handed curvature can also be formed by the coupling of the intrinsic tile curvature and the arm twist. We were able to assign the chiral indices (n,m) to each tube using high-resolution AFM images, and therefore were able to estimate the tile curvature using a regular polygon model that approximated the transverse section of the tube. A deeper understanding of the integrated actions of dif-ferent types of twisting forces on the DNA tubes will be extremely helpful in engineering more elaborate DNA nanostructures in the future.

A chiral dimer of an organic semiconductor was assembled from octaniline (octamer of polyaniline) conjugated to DNA. Facile reconfiguration between the monomer and dimer of octaniline-DNA was achieved. The geometry of the dimer and the exciton coupling between octaniline molecules in the assembly was studied both experimentally and theoretically. The octaniline dimer was readily switched between different electronic states by protonic doping and exhibited a Davydov splitting comparable to those previously reported for DNA-dye systems employing dyes with strong transition dipoles. This approach provides a possible platform for studying the fundamental properties of organic semiconductors with DNA-templated assemblies, which serve as candidates for artificial light-harvesting systems and excitonic devices.

p class="Text" style="margin: 0in 0in 8pt; text-align: justify; line-height: 24px; font-size: 12pt; font-family:quot;Times New Romanquot;, serif; color: rgb(0, 0, 0);"Non-canonical interactions in DNA remain under-explored in DNA nanotechnology. Recently, many structures with non-canonical motifs have been discovered, notably a hexagonal arrangement of typically rhombohedral DNA tensegrity triangles that forms through non-canonical sticky end interactions. Here, we find a series of mechanisms to program a hexagonal arrangement using: the sticky end sequence; triangle edge torsional stress; and crystallization condition. We showcase cross-talking between Watson-Crick and non-canonical sticky ends in which the ratio between the two dictates segregation by crystal forms or combination into composite crystals. Finally, we develop a method for reconfiguring the long-range geometry of formed crystals from rhombohedral to hexagonal anditalicvice versa/italic. These data demonstrate fine control over non-canonical motifs and their topological self-assembly. This will vastly increase the programmability, functionality, and versatility of rationally designed DNA constructs.o:p/o:p/p.

Chemical analysis of metalized DNA has made it quite clear that traditional models of DNA thermodynamics are insufficient to predict and control self-assembly in the context of orthogonally-paired nucleotides. Recently, there has been an increase in reports of Watson-Crick assembly of DNA wires and nanostructures [1–4]. The ability to add or remove pairing rules between nucleobases toward non-Watson-Crick, or orthogonal, self-assembly alters the fundamental language of DNA assembly: this change in behavior necessitates an accompanying shift in computational design. We begin by exploring the state-of-the-art in DNA modeling, and include both sequence analysis and sequence design practices. We then start from first principles and establish a mathematical basis for heterostructure and ‘nmer’ analysis in connected DNA networks that operates without assumptions about nucleobase parity. A generalized search algorithm is then constructed in Matlab and implemented using evolutionary techniques. We then discuss DNA nanostructure design criteria, operation efficiency in differentially-connected networks, and the application of computationally-aided sequence design for nanotechnological applications. We design a double crossover DNA motif with a silver base pair modification as a test case, and demonstrate successful model implementation. In sum, we present a novel computational framework for geometry-informed optimization of DNA networks. This tool is meant to enable design of both linear and nonlinear polynucleotide assemblies with inherent modularity for base parity, metalation, or more exotic nucleotide substitutions that may arise from advances in synthetic biology, nanomaterials and nanomedicine.

Colloidal synthesis is a powerful bottom-up approach for programmed self-assembly which holds promise for both research and industry. While diverse, each synthetic process is typically restricted to a specific chemistry. Many applications however require composite materials, whereas a chemical equilibrium can typically only match one material but not the other. Here, a scalable general approach is presented, alleviating the dependency on a specific chemical reaction, by resorting to a mechanical equilibrium; an isopycnic density-gradient-step is tailored to form clusters with prescribed composition. Valence control is demonstrated, making dimers, trimers, and tetramers with purity as high as 96%. The measured kinetics shows a scaleable throughput. The density gradient step plays a dual role of both filtering out undesired products and concentrating the target structures. The "Mix-and-Match" approach is general, and applies to a broad range of colloidal matter: diverse material compositions (plastics, glasses, and emulsions); a range of colloidal interactions (van der Waals, Coulomb, and DNA hybridization); and a spectrum of sizes (nanoscale to multiple micrometers). Finally, the strength of the method is displayed by producing a monodisperse suspension from a highly polydisperse emulsion. The ability to combine colloids into architectures of hybrid materials has applications in pharmaceuticals, cosmetics, and photonics.

DNA, when folded into nanostructures with a specific shape, is capable of spacing and arranging binding sites into a complex geometric pattern with nanometre precision. Here we demonstrate a designer DNA nanostructure that can act as a template to display multiple binding motifs with precise spatial pattern-recognition properties, and that this approach can confer exceptional sensing and potent viral inhibitory capabilities. A star-shaped DNA architecture, carrying five molecular beacon-like motifs, was constructed to display ten dengue envelope protein domain III (ED3)-targeting aptamers into a two-dimensional pattern precisely matching the spatial arrangement of ED3 clusters on the dengue (DENV) viral surface. The resulting multivalent interactions provide high DENV-binding avidity. We show that this structure is a potent viral inhibitor and that it can act as a sensor by including a fluorescent output to report binding. Our molecular-platform design strategy could be adapted to detect and combat other disease-causing pathogens by generating the requisite ligand patterns on customized DNA nanoarchitectures., DNA is capable of self-assembling into a wide range of user-defined structures and so can be used as a scaffold to arrange binding motifs with nanometre precision. Now, DNA has been used to accurately display aptamers that fit the repeated epitope pattern of a dengue viral antigen to produce a nanostructure that can be a potent viral inhibitor or a fluorescent sensor.

A quasi-one-dimensional organic semiconductor, hepta(p-phenylene vinylene) (HPV), was incorporated into a DNA tensegrity triangle motif using a covalent strategy. 3D arrays were self-assembled from an HPV-DNA pseudo-rhombohedron edge by rational design and characterized by X-ray diffraction. Templated by the DNA motif, HPV molecules exist as single-molecule fluorescence emitters at the concentration of 8 mM within the crystal lattice. The anisotropic fluorescence emission from HPV-DNA crystals indicates HPV molecules are well aligned in the macroscopic 3D DNA lattices. Sophisticated nanodevices and functional materials constructed from DNA can be developed from this strategy by addressing functional components with molecular accuracy.

The successful self-assembly of tensegrity triangle DNA crystals heralded the ability to programmably construct macroscopic crystalline nanomaterials from rationally-designed, nanoscale components. This 3D DNA tile owes its "tensegrity" nature to its three rotationally stacked double helices locked together by the tensile winding of a center strand segmented into 7 base pair (bp) inter-junction regions, corresponding to two-thirds of a helical turn of DNA. All reported tensegrity triangles to date have employed

The rational design of nanoscopic DNA tiles has yielded highly ordered crystalline matter in 2D and 3D. The most well-studied 3D tile is the DNA tensegrity triangle, which is known to self-assemble into macroscopic crystals. However, contemporary rational design parameters for 3D DNA crystals nearly universally invoke integer numbers of DNA helical turns and Watson-Crick (WC) base pairs. In this study, 24-bp edges are substituted into a previously 21-bp (two helical turns of DNA) tensegrity triangle motif to explore whether such unconventional motif can self-assemble into 3D crystals. The use of noncanonical base pairs in the sticky ends results in a cubic arrangement of tensegrity triangles with exceedingly high symmetry, assembling a lattice from winding helical axes and diamond-like tessellation patterns. Reverting this motif to sticky ends with Watson-Crick pairs results in a trigonal hexagonal arrangement, replicating this diamond arrangement in a hexagonal context. These results showcase that the authors can generate unexpected, highly complex, pathways for materials design by testing modifications to 3D tiles without prior knowledge of the ensuing symmetry. This study expands the rational design toolbox for DNA nanotechnology; and it further illustrates the existence of yet-unexplored arrangements of crystalline soft matter.

The tensegrity triangle motif utilizes Watson-Crick sticky end cohesion to self-assemble into a rhombohedral crystal lattice using complementary 5'-GA and 5'-TC sticky ends. Here, we report that using noncanonical 5'-AG and 5'-TC sticky ends in otherwise isomorphic tensegrity triangles results in crystal self-assembly in the P63 hexagonal space group as revealed by X-ray crystallography. In this structure, the DNA double helices bend at the crossover positions, a feature that was not observed in the original design. Instead of propagating linearly, the tilt between base pairs of each right-handed helix results in a left-handed superstructure along the screw axis, forming a microtubule-like structure composed of three double helices with an unbroken channel at the center. This hexagonal lattice has a cavity diameter of 11 nm and a unit cell volume of 886 000 A3-far larger than the rhombohedral counterpart (5 nm, 330 000 A3).

Sticky-end cohesion plays a critical role in molecular biology and nucleic acid nanotechnology. Although free energy calculations and molecular mechanics can predict these interactions, chemical modification would compromise such predictions. Herein, we have used rationally designed 3D DNA crystals as a tool to experimentally investigate the modulation of 5'-phosphorylation on sticky-end cohesions. We have found that 5'-phosphorylation strengthens the sticky-end cohesion: in a DNA crystal self-assembled exclusively via sticky-end cohesions, 5'-phosphorylation not only promotes the crystallization process, in general, but also accelerates the crystal growth along designed directions. Such a finding allows the fine-tuning of DNA crystallization kinetics and the control of DNA crystal morphology. It also suggests a potential difference in self-assembly kinetics between natural DNA (with 5'-phosphorylation) and synthetic DNA (without 5'-phosphorylation).

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. The first one is based on immobile Holliday junctions which employ specific nucleotide sequences at the branch points which dictate their orientation. The second strategy is to use angle-enforcing struts to fix the branching orientation with flexible spacers at the branch points. We have also demonstrated that the branching orientation control can be achieved dynamically, either by canonical Watson-Crick base pairing or non-canonical nucleobase interactions ( e.g., i-motif and G-quadruplex). With precise angle control and feedback from the chemical environment, these results will enable novel DNA nanomechanical sensing devices, and precisely-ordered three-dimensional architectures.

Two-dimensional (2D) DNA origami that is capable of self-assembling into complex 2D and 3D geometries pave the way for a bottom-up synthesis for various applications in nano/biotechnology. Here, we directly visualized the aqueous structure of 2D DNA origami cross-tiles and their assemblies using cryogenic electron microscopy. We uncovered flexible arms in cross-tile monomers and designated inter-tile folding. In addition, we observed the formation of clusters and stacks of DNA cross-tiles in solution, which could potentially affect the interaction and assembly of DNA origami. Finally, we quantitatively evaluated the flexibility of DNA origami in solution using finite element analysis. Our discovery has laid the foundation for investigating the dynamic structures of 2D DNA origami assemblies in solution, providing insights regarding the self-assembly and self-replication mechanisms of 2D DNA origami.

DNA overwinding and underwinding between adjacent Holliday junctions have been applied in DNA origami constructs to design both left-handed and right-handed nanostructures. For a variety of DNA tubes assembled from small tiles, only a theoretical approach of the intrinsic tile curvature was previously used to explain their formation. Details regarding the quantitative and structural descriptions of the intrinsic tile curvature and its evolution in DNA tubes by coupling with arm twists were missing. In this work, we designed three types of tile cores from a circular 128 nucleotide scaffold by longitudinal weaving (LW), bridging longitudinal weaving (bLW), and transverse weaving (TW) and assembled their 2D planar or tubular nanostructures via inter-tile arms with a distance of an odd or even number of DNA half-turns. The biotin/streptavidin (SA) labeling technique was applied to define the tube configuration with addressable inside and outside surfaces and thus their component tile conformation with addressable concave and convex curvatures. Both chiral tubes possessing left-handed and right-handed curvatures could be generated by finely tuning p and q in bLW-Ep/q designs (bLW tile cores joined together by inter-tile arms of an even number of half-turns with the arm length of p base pairs (bp) and the sticky end length of q nucleotides (nt)). We were able to assign the chiral indices (n,m) to each specific tube from the high-resolution AFM images, and thus estimated the tile curvature angle with a regular polygon model that approximates each tube’s transverse section. We attribute the curvature evolution of bLW-Ep/q tubes composed of the same tile core to the coupling of the intrinsic tile curvature and different arm twists. A better understanding of the integrated actions of different types of twisting forces on DNA tubes will be much more helpful in engineering DNA nanostructures in the future.

Significance With simple DNA origami lever arms arranged in hinges and accordion structures, we amplify the nanometer displacements from DNA hairpin zippers to 4-μm motion, easily observable and quantified in real space and real time with conventional optical microscopy. Mechanically pulling a bead tethered on the accordion end, we measure high-energy recovery and retraction speeds up to 50 μm/s. On longer time scales, we have also opened and closed the hinges with light and heat. DNA nanotechnology, and particularly DNA origami, combined with colloids and emulsions can provide powerful architectures. The present study is a step toward activating such colloidal/cellular scale devices using DNA as a power source/fuel. We envision artificial active flagella, cilia, micropumps, and other cellular scale devices., The programmability of DNA oligonucleotides has led to sophisticated DNA nanotechnology and considerable research on DNA nanomachines powered by DNA hybridization. Here, we investigate an extension of this technology to the micrometer-colloidal scale, in which observations and measurements can be made in real time/space using optical microscopy and holographic optical tweezers. We use semirigid DNA origami structures, hinges with mechanical advantage, self-assembled into a nine-hinge, accordion-like chemomechanical device, with one end anchored to a substrate and a colloidal bead attached to the other end. Pulling the bead converts the mechanical energy into chemical energy stored by unzipping the DNA that bridges the hinge. Releasing the bead returns this energy in rapid (>20 μm/s) motion of the bead. Force-extension curves yield energy storage/retrieval in these devices that is very high. We also demonstrate remote activation and sensing—pulling the bead enables binding at a distant site. This work opens the door to easily designed and constructed micromechanical devices that bridge the molecular and colloidal/cellular scales.

Engineered 3D DNA crystals are promising scaffolds for bottom-up construction of three-dimensional, macroscopic devices from the molecular level. Nevertheless, this has been hindered by the highly constrained conditions for DNA crystals to be stable. Here we report a method to prepare robust 3D DNA crystals by postassembly ligation to remove this constraint. Specifically, sticky ends at crystal contacts were enzymatically ligated, and the covalent bonds significantly enhanced crystal stability, e.g., being stable at 65 °C. This method also enabled the fabrication of DNA crystals with complex architectures including crystal shell, core-shell, and matryoshka dolls. Furthermore, we have demonstrated the applications of the robust DNA crystals in biocatalysis and protein entrapment. Our study removes one key obstacle for the applications of DNA crystals and offers many new opportunities in DNA nanotechnology.