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A novel supramolecular DNA hydrogel system was designed based on a directly synthesized chemically branched DNA. For the hydrogel formation, a self-dimer DNA with two sticky ends was designed as the linker to induce the gelation of B-Y. By programing the linker sequence, thermal and metal-ion responsiveness could be introduced into this hydrogel system. This supramolecular DNA hydrogel shows shear-thinning, designable responsiveness, and good biocompatibility, which will simplify the hydrogel composition and preparation process of the supramolecular DNA hydrogel and accelerate its biomedical applications.

The chemical interactions that lead to the emergence of hierarchical structures are often highly complex and difficult to program. Herein, the synthesis of a series of superlattices based upon 30 different structurally reconfigurable DNA dendrimers is reported, each of which presents a well-defined number of single-stranded oligonucleotides (i.e., sticky ends) on its surface. Such building blocks assemble with complementary DNA-functionalized gold nanoparticles (AuNPs) to yield five distinct crystal structures, depending upon choice of dendrimer and defined by phase symmetry. These DNA dendrimers can associate to form micelle-dendrimers, whereby the extent of association can be modulated based upon surfactant concentration and dendrimer length to produce a low-symmetry Ti5Ga4-type phase that has yet to be reported in the field of colloidal crystal engineering. Taken together, colloidal crystals that feature three different types of particle bonding interactions-template-dendron, dendrimer-dendrimer, and DNA-modified AuNP-dendrimer-are reported, illustrating how sequence-defined recognition and dynamic association can be combined to yield complex hierarchical materials.

Annealing between sticky ends of DNA is an intermediate step in ligation. It can also be utilized to program specific binding sites for DNA tile and origami assembly. This reaction is generally understood as a bimolecular reaction dictated by the local concentration of the sticky ends. Its dependence on the relative orientation between the sticky ends, however, is less understood. Here we report on the interactions between DNA sticky ends using the coarse-grained oxDNA model; specifically, we consider how the orientational alignment of the double-stranded DNA (dsDNA) segments affects the time required for the sticky ends to bind, τ(b). We specify the orientation of the dsDNA segments with three parameters: θ, which measures the angle between the helical axes, and ϕ(1) and ϕ(2), which measure rotations of each strand around the helical axis. We find that the binding time depends strongly on both θ and ϕ(2): ~20-fold change with θ and 10-fold change with ϕ(2). The binding time is the fastest when the helical axes of duplexes are pointing toward each other and the sticky ends protrude from the farthest two points. Our result is relevant for predicting hybridization efficiency of sticky ends that are rotationally restricted.

We report a new DNA nanostructure, an extended 1-dimensional composite built for the first time out of structurally robust yet conveniently disassembled DNA triple helices, interspersed with short stretches of G-quadruplexes. These "TQ Hybrid" 1-dimensional nanostructures require potassium ions and modestly acidic pH for their formation and are easily disassembled by changes to either of these requirements. We initially prepared and characterized a "monomeric" TQ Hybrid tile; followed by "sticky" TQs tiles, incorporating unique guanine-only sticky ends, that enable efficient self-assembly via G-quartet formation of nanostructures >150 nm in length, as seen with atomic force microscopy and transmission electron microscopy. We anticipate that such DNA TQ Hybrid structures will find unique and varied application as communication modules within larger nanostructures, and as sensors, logic gates, as well as in other aspects of DNA nanotechnology.

Phase separation of biomolecules plays key roles in physiological compartmentalization as well as pathological aggregation. A deeper understanding of biomolecular phase separation requires dissection of a relation between intermolecular interactions and resulting phase behaviors. DNA nanostars, multivalent DNA assemblies of which sticky ends define attractive interactions, represent an ideal system to probe this fundamental relation governing phase separation processes. Here, we use DNA nanostars to systematically study how structural flexibility exhibited by interacting species impacts their phase behaviors. We design multiple nanostars with a varying degree of flexibility using single-stranded gaps of different lengths in the arm of each nanostar unit. We find that structural flexibility drastically alters the phase diagram of DNA nanostars in such a way that the phase separation of more flexible structures is strongly inhibited. This result is not due to self-inhibition from the loss of valency but rather ascribed to a generic flexibility-driven change in the thermodynamics of the system. Our work provides not only potential regulatory mechanisms cells may exploit to dynamically control intracellular phase separation but also a route to build synthetic systems of which assembly can be controlled in a signal dependent manner.

A key challenge for the discrete distribution-based Förster resonance energy transfer system (D-FRET) is the reduced intensity and stability of signal probes in complex biological matrices. Here, we present a spatially confined FRET (SC-FRET) probe with a stable structure and strong signal output. It consists of multivalent FRET pairs labeled with FAM or TAMRA. In this assay, p53 DNA was chosen as a model hairpin probe (HP), and two kinds of branched DNA probes (ssDNA-FAM, ssDNA-TAMRA) were involved. Under the action of p53 DNA, the unfolded HP acts as a primer to initiate polymerization extension of KFP polymerase and cleavage of Nb.BbvCI endonuclease, which produces plenty of ssDNA (primer-DNA). The branched DNA is designed to have the same binding core and different sticky ends, the core part of which can self-assemble to form X-shaped branched DNA (X-FAM or X-TAMRA), and the sticky ends of which are complementary to the primer-DNA. Therefore, the primer-DNAs released during the polymerization cleavage process will combine a large number of X-FAM and X-TAMRA in a limited space through complementary base pairing. Fluorescence was transferred from FAM to TAMRA, and a strong FRET response was generated by the locational effects. The proposed SC-FRET system based on the multivalent assembly of branched DNA exhibited a strong FRET response with an LOD of 0.01394 pM. Importantly, it also showed a high-contrast and stable FRET response in HeLa cells. Its superior biological stability is attributed to the large steric hindrance of the compact and rigid frame of the SC-FRET probe, which helps prevent intracellular degradation and provides a powerful tool for biomedical research.

Telomeres are associated with the ends of DNA double strands. The lengths of the telomeres are controlled by the telomerase enzyme. The shortening of the telomeres is known to relate to aging. In cancers, telomere lengths are abnormally short. Telomeres could act as buffers shielding the part of DNA coding for the proteins. For cancer cells, germ cells and stem cells the length of the telomeres is not varying. There is an analogy with microtubules, which are highly dynamical and carry a longitudinal electric field, whose strength correlates with the microtubule length. Could sticky ends generate a longitudinal field along DNA double strand with strength determined by the lengths of the sticky ends? In the standard picture the flux of the longitudinal electric field would be proportional to the difference of the negative charges associated with the sticky ends. In TGD framework, DNA strands are accompanied by the dark analog of DNA with codons realized as 3-proton units at magnetic flux tubes parallel to DNA strands and neutralizing the negative charge of ordinary DNA except at the sticky ends. This allows considering the possibility that opposite sticky ends carry opposite charges generating a longitudinal electric field along the magnetic flux tube associated with the system. DNA/Telomere bioelectric field could serve as a novel bioelectric marker to be used for prognostic and diagnostic purposes in researches of cancer, aging, surgery grafts and rejuvenation. We propsed that DNA bioelectric field can be used as a futuristic bioelectric marker of cancer, aging and death.

Target-oriented cellular automata with computation are the primary challenge in the field of DNA algorithmic self-assembly in connection with specific rules. We investigate the feasibility of using the principle of cellular automata for mathematical subjects by using specific logic gates that can be implemented into DNA building blocks. Here, we connect the following five representative elementary functions: (i) enumeration of multiples of 2, 3, and 4 (demonstrated via R094, R062, and R190 in 3-input/1-output logic rules); (ii) the remainder of 0 and 1 (R132); (iii) powers of 2 (R129); (iv) ceiling function for n/2 and n/4 (R152 and R144); and (v) analogous pattern of annihilation (R184) to DNA algorithmic patterns formed by specific rules. After designing the abstract building blocks and simulating the generation of algorithmic lattices, we conducted an experiment as follows: designing of DNA tiles with specific sticky ends, construction of DNA lattices via a two-step annealing method, and verification of expected algorithmic patterns on a given DNA lattice using an atomic force microscope (AFM). We observed representative patterns, such as horizontal and diagonal stripes and embedded triangles, on the given algorithmic lattices. The average error rates of individual rules are in the range of 8.8% (R184) to 11.9% (R062), and the average error rate for all the rules was 10.6%. Interpretation of elementary functions demonstrated through DNA algorithmic patterns could be extended to more complicated functions, which may lead to new insights for achieving the final answers of functions with experimentally obtained patterns.

Extensive manipulations involved in the preparation of DNA samples for sequencing have hitherto made it impossible to determine the precise structure of double-stranded DNA fragments being sequenced, such as the presence of blunt ends, single-stranded overhangs, or single-strand breaks. We here describe MatchSeq, a method that combines single-stranded DNA library preparation from diluted DNA samples with computational sequence matching, allowing the reconstruction of double-stranded DNA fragments on a single-molecule level. The application of MatchSeq to Neanderthal DNA, a particularly complex source of degraded DNA, reveals that 1- or 2-nt overhangs and blunt ends dominate the ends of ancient DNA molecules and that short gaps exist, which are predominantly caused by the loss of individual purines. We further show that deamination of cytosine to uracil occurs in both single- and double-stranded contexts close to the ends of molecules, and that single-stranded parts of DNA fragments are enriched in pyrimidines. MatchSeq provides unprecedented resolution for interrogating the structures of fragmented double-stranded DNA and can be applied to fragmented double-stranded DNA isolated from any biological source. The method relies on well-established laboratory techniques and can easily be integrated into routine data generation. This possibility is shown by the successful reconstruction of double-stranded DNA fragments from previously published single-stranded sequence data, allowing a more comprehensive characterization of the biochemical properties not only of ancient DNA but also of cell-free DNA from human blood plasma, a clinically relevant marker for the diagnosis and monitoring of disease.

DNA is an ideal molecule for the construction of 3D crystals with tunable properties owing to its high programmability based on canonical Watson-Crick base pairing, with crystal assembly in all three dimensions facilitated by immobile Holliday junctions and sticky end cohesion. Despite the promise of these systems, only a handful of unique crystal scaffolds have been reported. Herein, we describe a new crystal system with a repeating sequence that mediates the assembly of a 3D scaffold via a series of Holliday junctions linked together with complementary sticky ends. By using an optimized junction sequence, we could determine a high-resolution (2.7 Å) structure containing R3 crystal symmetry, with a slight subsequent improvement (2.6 Å) using a modified sticky-end sequence. The immobile Holliday junction sequence allowed us to produce crystals that provided unprecedented atomic detail. In addition, we expanded the crystal cavities by 50 % by adding an additional helical turn between junctions, and we solved the structure to 4.5 Å resolution by molecular replacement.

We use computer simulations and experiments to study the interactions between nanoparticles (NPs) grafted with self-complementary DNA strands. Each strand ends with a sticky palindromic single-stranded sequence, allowing it to associate equally favorably with strands grafted on the same particle or on different NPs. Surprisingly we find an attractive interaction between a pair of NPs, and we demonstrate that at low temperature it arises purely from a combinatorial-entropy contribution. We evaluate theoretically and verify numerically this entropic contribution originating from the number of distinct bonding patterns associated with intra- and interparticle binding. This entropic attraction becomes more favorable with decreasing inter-NP distance because more sticky ends can participate in making this choice.

Cyclization of DNA with sticky ends is commonly used to measure DNA bendability as a function of length and sequence, but how its kinetics depend on the rotational positioning of the sticky ends around the helical axis is less clear. Here, we measured cyclization (looping) and decyclization (unlooping) rates (kloop and kunloop) of DNA with sticky ends over three helical periods (100-130 bp) using single-molecule fluorescence resonance energy transfer (FRET). kloop showed a nontrivial undulation as a function of DNA length whereas kunloop showed a clear oscillation with a period close to the helical turn of DNA (∼10.5 bp). The oscillation of kunloop was almost completely suppressed in the presence of gaps around the sticky ends. We explain these findings by modeling double-helical DNA as a twisted wormlike chain with a finite width, intrinsic curvature, and stacking interaction between the end base pairs. We also discuss technical issues for converting the FRET-based cyclization/decyclization rates to an equilibrium quantity known as the J factor that is widely used to characterize DNA bending mechanics.

A rapid and colorimetric biosensor for Pb2+ detection has been constructed on the basis of Pb2+-dependent GR-5 DNAzyme and the self-replicating catalyzed hairpin assembly (SRCHA) reaction. In the presence of Pb2+, the phosphodiester bond of the internal RNA base (rA) in GR-5 DNAzyme was cleaved by the enzyme strand and trigger DNA was released. Then the SRCHA reaction was initiated and the H1–H2 complex equipped with single-stranded DNA (ssDNA) sticky ends on both sides was formed. The sequences of the newly formed sticky ends on one side of H1–H2 were the same as that of trigger DNA and then additional CHA reaction cycles were initiated. Meanwhile, the intact G-quadruplex was reunited on the other side of H1–H2 for colorimetric signal readout. By taking advantage of the self-replication of trigger DNA, the response signal in this sensing system was significantly amplified and Pb2+ can be detected in a linear range from 20 to 100 nM with a detection limit of 2.6 nM within 20 min. In addition, this method can be applied to the reliable monitoring of spiked Pb2+ in environmental water and serum samples with satisfying results, providing a potential method for on-site and real-time detection of Pb2+ for environmental protection and related disease prevention.

Thermus thermophilus DNA ligase (Tth DNA ligase) is widely employed for cloning, enzymatic synthesis, and molecular diagnostics at high temperatures (e.g., 65 °C). It has been long believed that the complementary ends must be very long (e.g., >30 bp) to place two DNA fragments nearby for the ligation. In the current study, the length of the complementary portion was systematically varied, and the ligation efficiency was evaluated using the high resolution melting (HRM) method. Unexpectedly, very short oligonucleotides (7-10 nt) were successfully ligated on the complementary overhang attached to a dsDNA at 70 °C. Furthermore, sticky ends with the overhang of only 4 nt long, available after scission with many restriction enzymes, were also efficiently ligated at 45-70 °C. The ligation yield for the 6-nt-long sticky ends was as high as 80%. It was concluded that Tth DNA ligase can be used as a unique tool for DNA manipulation that cannot be otherwise easily accomplished.

MicroRNAs (miRNAs) are found in extremely low concentrations in cells, so highly sensitive quantitation is a great challenge. Herein, a simple dual-amplification strategy involving target-activated catalytic hairpin assembly (CHA) coupled with multiple fluorophores concentrated on one X-shaped DNA is reported. In this strategy, four hairpin probes (H1, H2, H3, and H4) are modified with FAM and BHQ1 at both sticky ends, while a circulating hairpin probe (H0) is used to activate CHA circuits once it binds to complementary sequences in the target miR-21 (T). The powerful dual-amplification cascades in Forster resonance energy transfer (FRET)-based nonenzymatic nucleic acid circuits are triggered by T-H0-activated formation of the X-shaped DNA nanostructure, freeing T-H0 for the next CHA reaction cycle. CHA circuits increase the fluorescence due to the wide distance between FAM and BHQ1 in the formed X-shaped DNA nanostructure, resulting in signal amplification and highly sensitive detection of miR-21, with a limit of detection (LOD, 3σ) of 0.025 nM, which is 25.6 or 57.6 times lower than that obtained through a single-amplification strategy without multiple fluorophores on one X-shaped DNA or CHA circuit. Furthermore, this cascade reaction was completed in 45 min, effectively avoiding target degradation. This new enzyme-free signal amplification strategy holds promising potential for sensitively detecting different DNA or RNA sequences by simply adapting the fragment of the H0 sequence complementary to the target.

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).

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.

DNA tensegrity triangles self-assemble into rhombohedral three-dimensional crystals via sticky ended cohesion. Crystals containing two-nucleotide (nt) sticky ends (GA:TC) have been reported previously, and those crystals diffracted to 4.9 Å at beam line NSLS-I-X25. Here, we analyze the effect of varying sticky end lengths and sequences, as well as the impact of 5’- and 3’-phosphates on crystal formation and resolution. Tensegrity triangle motifs having 1-, 2- and 3-nt sticky ends all form crystals. X-ray diffraction data from the same beam line reveal that the crystal resolution for a 1-nt sticky end (G:C) and a 3-nt sticky end (GAT:ATC) were 3.4 Å and 4.2 Å respectively. Resolutions were determined from complete data sets in each case. We also conducted trials that examined every possible combination of 1-nucleotide and 2-nucleotide sticky-ended phosphorylated strands and successfully crystallized all 16 possible combinations of strands. We observed the position of the 5’-phosphate on either the crossover (1), helical (2), or central strand (3) affected the resolution of the self-assembled crystals for the 2-turn monomer (3.0 Å for 1–2P-3P) and 2-turn dimer sticky ended (4.1 Å for 1–2-3P) systems. We have also examined the impact of the identity of the base flanking the sticky ends, as well as the use of 3’-sticky ends. We conclude that crystal resolution is not a simple consequence of the thermodynamics of the direct nucleotide pairing interactions involved in molecular cohesion in this system.

Molecular-genetic research methods make it possible to identify phytopathogens with comprehensive characteristics of their hereditary material. We suggest a genotyping method based on the use of two restriction endonucleases for the cleavage of bacteria’s genomic DNA. Taq DNA polymerase added to the reaction mixture provides labeling of DNA fragments with biotinylated deoxycytosinetriphosphate (Bio-dCTP). The label is only incorporated into DNA having 3-prime recessed ends formed by the first enzyme. The second restriction endonuclease produces only blunt ends of fragments that are unable to incorporate the label. As a result of the DDSL (double digest selective label) reaction, 20–50 distinct DNA fragments are visualized on the filter, the amount and distribution of which are specific for each bacterial strain. This research was carried out using two pairs of restriction enzymes (XbaI/DraI and XbaI/Eco24I). The results indicate the same discriminatory capacity of these enzymes. Some advantage was noted for the XbaI/DraI combination, which makes it possible to detect differences in the region of longer DNA fragments in genetic profiles. The genetic profile generated by the BcuI/Eco32I pair of enzymes in Ps. fluorescens 894 was significantly different from that of Ps. xanthochlora, which could be anticipated while comparing these two species. In general, the results of genotyping agreed with the data on the origin of bacterial strains. In particular, microorganisms with the same genetic profile, D822 and D828, G399 and G400, as well as C960 and C966, were obtained from affected tubers of the same experimental field. This suggests the possibility of the contact spread of the pathogen. In some cases, the identity of strains was observed, despite their different geographic origin. This fact confirms contamination that occurred in the past during the use of the foreign seed potato.

The photosensitive azobenzene-tethered DNA material provides an ideal force for DNA nanomachines. This work demonstrates the process of light-driven DNA strand displacement by physical and mathematical models. By data fitting, the strand displacement reaction rate constants under different sequence designs are discussed in detail. The results showed that the Gibbs free energy induced by azobenzene is around 2.52 kJ mol−1, which would hinder the light-driven strand displacement reaction. Increasing the length of the sticky ends can increase the displacement rate exponentially, but the displacement rate will not increase when it exceeds 12 bases.

Solution-phase self-assembly of DNA-functionalized nanoparticles into mesoscale structures is a promising strategy for creating functional materials from nanocrystal building blocks. The predominant approach has been the use of Watson-Crick base pairing between complementary bases in designated 'sticky ends' to trigger programmable self-assembly into ordered superlattices. Here we demonstrate the ordered self-assembly of Au nanoparticles conjugated with single-stranded (ss) DNA in acidic solutions. Au nanoparticles functionalized with thiolated ssDNA are protected against coalescence and the DNA conformation undergoes significant modifications at low pH, which can be associated with the protonation of adenine bases and the formation of a parallel poly-adenine duplex, which govern the interaction between ssDNA-Au nanoparticle conjugates. In situ liquid cell electron microscopy enables real-time imaging of the self-assembly process and the identification of key characteristics, such as the preferred structural motifs and interparticle separations in the native solution environment. Our results highlight alternatives to conventional base-pairing interactions for building DNA-directed nanoparticle superlattices.

A novel and rapid room-temperature DNA amplification strategy to simultaneously amplify and detect target HIV DNA was developed by coupling catalyzed hairpin assembly (CHA) with a “non-sequence dependent” nicking endonuclease. In this system, a Nt.BsmAI nicking endonuclease recognition sequence and HIV target DNA sequence were integrated into hairpin probes H1 and H3, respectively. The HIV target DNA was repeatedly used to trigger a CHA reaction, forming numerous H1–H2 duplexes. Then the newly formed sticky ends of H1–H2 caused the hairpin structure of H3 to open. As a result, the double-stranded nicking enzyme recognition site was reunited, which was recognized and cleaved by the nicking enzyme. Finally, the target DNA replica with a sequence matching that of the HIV target DNA was obtained. The above reactions, namely, the CHA reaction, hairpin opening and enzyme cleavage, can be continuously repeated, leading to rapid room-temperature DNA amplification. In addition, this proposed rapid DNA amplification strategy enables picomolar detection of HIV DNA target within 10 min, providing a potential clinical application at point-of-care.

In typical photoelectrochemical (PEC) biosensing assays, electrodes are generally modified with photoactive probes and/or target recognition probes, which makes the processes complicated, time-consuming, and difficult to achieve excellent reproducibility. Hence, to overcome such shortcomings, we propose here an immobilization-free and label-free PEC aptasensor using solution-phase methylene blue (MB) as the PEC signal probe. Based on the unique T-Hg2+-T base pairs, and the diffusivity difference between free MB molecules and the MB/G-quadruplex composite towards the ITO electrode surface with negative charge, the "signal-off" approach for Hg2+ detection is developed. In the presence of target Hg2+, via the T-Hg2+-T bond formation, the two sticky ends of the hairpin DNA probe form a rigid duplex stem, which triggers the exonuclease III-facilitated target cycling amplification, and the formation of multiple G-quadruplexes. Upon the intercalation of MB in G-quadruplexes, significantly decreased photocurrent is obtained owing to the increased electrostatic repulsion between the MB/G-quadruplex composite and the ITO electrode. Therefore, highly sensitive and ultrasensitive Hg2+ determination is achieved, with a low detection limit of 1.2 pM, well below the maximum allowable Hg2+ level in drinking water defined by the WHO, China's Ministry of Health, and the US EPA. Due to the avoidance of sophisticated electrode modification and recognition probe immobilization processes, as well as an expensive labeling procedure, the PEC aptasensor proposed here demonstrates the advantages of simplicity, good reproducibility, rapidness and low cost.

Nature uses non-covalent interactions to achieve structural dynamic reconfiguration of biopolymers. Taking advantage of the programmability of DNA/DNA interactions we report here the rational design of orthogonal DNA-based addressable tiles that self-assemble into polymer-like structures that can be reconfigured by external inputs. The different tiles share the same sticky ends responsible for self-assembly but are rationally designed to contain a specific regulator-binding domain that can be orthogonally targeted by different DNA regulator strands. We show that by sequentially adding specific inputs it is possible to re-organize the formed structures to display well-defined distributions: homopolymers, random and block structures. The versatility of the systems presented in this study shows the ease with which DNA-based addressable monomers can be designed to create reconfigurable micron-scale DNA structures offering a new approach to the growing field of supramolecular polymers.

Oligonucleotide-functionalized nanoparticles (NPs), also known as "programmable atom equivalents" (PAEs), have emerged as a class of versatile building blocks for generating colloidal crystals with tailorable structures and properties. Recent studies have shown that, at small size and low DNA grafting density, PAEs can also behave as "electron equivalents" (EEs), roaming through and stabilizing a complementary PAE sublattice. However, it has been challenging to obtain a detailed understanding of EE-PAE interactions and the underlying colloidal metallicity because there is inherent polydispersity in the number of DNA strands on the surfaces of these NPs; thus, the structural uniformity and tailorability of NP-based EEs are somewhat limited. Herein, we report a strategy for synthesizing colloidal crystals where the EEs are templated by small molecules, instead of NPs, and functionalized with a precise number of DNA strands. When these molecularly precise EEs are assembled with complementary NP-based PAEs, X-ray scattering and electron microscopy reveal the formation of three distinct "metallic" phases. Importantly, we show that the thermal stability of these crystals is dependent on the number of sticky ends per EE, while lattice symmetry is controlled by the number and orientation of EE sticky ends on the PAEs. Taken together, this work introduces the notion that, unlike conventional electrons, EEs that are molecular in origin can have a defined valency that can be used to influence and guide specific phase formation.

Coating colloidal particles with DNA provides one of the most versatile and powerful methods for controlling colloidal self-assembly. Previous studies have shown how combining DNA coatings with DNA strand displacement allows one to design phase transitions between different three-dimensional crystal structures. Here we show that by using DNA coatings with bifunctional colloidal Janus particles, we can realize reconfigurable thermally reversible transitions between one- and two-dimensional self-assembled colloidal structures. We introduce a colloidal system in which DNA-coated asymmetric Janus particles can reversibly switch their Janus balance in response to temperature, resulting in the reconfiguration of assembling structures between colloidal chains and bilayers. Each face of the Janus particles is coated with different self-complementary DNA strands. Toehold strand displacement is employed to selectively activate or deactivate the sticky ends on the smaller face, which enables Janus particles to selectively assemble through either the smaller or larger face. This strategy could be useful for constructing complex systems that could be reconfigured to assemble into different structures in different environments.

Non-homologous end joining (NHEJ) is a highly conserved mechanism of DNA double-stranded break (DSB) repair. Here we utilize a computational protein-protein interaction method to identify human PRKACB as a potential candidate interacting with NHEJ proteins. We show that the deletion of its yeast homolog, TPK1 that codes for the protein kinase A catalytic subunit reduces the efficiency of NHEJ repair of breaks with overhangs and blunt ends in plasmid-based repair assays. Additionally, tpk1Δ mutants showed defects in the repair of chromosomal breaks induced by HO-site specific endonuclease. Our double deletion mutant analyses suggest that TPK1 and YKU80, a key player in NHEJ could function in parallel pathways. Altogether, here we report a novel involvement for TPK1 in NHEJ.

MicroRNAs (miRNAs) have been shown to be promising biomarkers for disease diagnostics and therapeutics. However, the rapid, low-cost, sensitive, and selective detection of miRNAs remains a challenge because of their characters of small size, vulnerability to degradation, low abundance, and sequence similarity. Herein, we describe an enzyme-free amplification platform, consisting of a catalytic hairpin assembly (CHA) and DNA-templated silver nanoclusters (DNA/AgNCs), for miRNA analysis. In this work, two DNA hairpins (H1 and H2) were first designed for target miR-21-induced CHA, and then the fluorescence of DNA/AgNCs was quenched by BHQ1 to construct an activatable probe (AP). In the presence of target miR-21, hairpin H1 was opened by miR-21 through a hybridization reaction, and hairpin H2 was then opened by H1. During this process, miR-21 was released from H1 and participated in the next round of hybridization, triggering the CHA cycle reaction. The obtained H1-H2 products with sticky ends could react with the AP, forcing BHQ1 away from the DNA/AgNCs and thus causing the fluorescence recovery of the DNA/AgNCs. The assay for miR-21 detection demonstrated an excellent linear response to concentrations varying from 200 pM to 20 nM with the detection limit of 200 pM. The simple and cost-effective strategy holds great potential for application in biomedical research and clinical diagnostics.

Membrane-active cytoskeletal elements, such as FtsZ, septin or actin, form filamentous polymers able to induce and stabilize curvature on cellular membranes. In order to emulate the characteristic dynamic self-assembly properties of cytoskeletal subunits in vitro, biomimetic synthetic scaffolds were here developed using DNA origami. In contrast to our earlier work with pre-curved scaffolds, we specifically assessed the potential of origami mimicking straight filaments, such as actin and microtubules, by origami presenting cholesteryl anchors for membrane binding and additional blunt end stacking interactions for controllable polymerization into linear filaments. By assessing the interaction of our DNA nanostructures with model membranes using fluorescence microscopy, we show that filaments can be formed, upon increasing MgCl2 in solution, for structures displaying blunt ends; and can subsequently depolymerize, by decreasing the concentration of MgCl2. Distinctive spike-like membrane protrusions were generated on giant unilamellar vesicles at high membrane-bound filament densities, and the presence of such deformations was reversible and shown to correlate with the MgCl2-triggered polymerization of DNA origami subunits into filamentous aggregates. In the end, our approach reveals the formation of membrane-bound filaments as a minimal requirement for membrane shaping by straight cytoskeletal-like objects.

© 2020 American Chemical Society. The self-assembly of collagen-mimetic peptides (CMPs) that form sticky-ended triple helices has allowed the production of surprisingly stable artificial collagen fibers and hydrogels. Assembly through sticky ends requires the recognition of a single strand by a templated strand dimer. Although CMPs and their triple helices have been studied extensively, the structure of a strand dimer is unknown. Here, we evaluate the physical characteristics of such dimers, using disulfide-templated (PPG)10 dimers as a model. Such "linked-dimers" retain their collagen-like structure even in the absence of a third strand, but only when their strands are capable of adopting a triple-helical fold. The intrinsic collagen-like structure of templated CMP pairs helps to explain the success of sticky-ended CMP association and changes the conception of new synthetic collagen designs.

The self-assembly of DNA hybrids possessing tetraphenylethylene sticky ends at both sides into vesicular architectures in aqueous medium is demonstrated. Cryo-electron microscopy reveals the formation of different types of morphologies from the amphiphilic DNA-hybrids. Depending on the conditions, either an extended (sheet-like) or a compact (columnar) alignment of the DNA hybrids is observed. The different modes of DNA arrangement lead to the formation of vesicles appearing either as prolate ellipsoids (type I) or as spheres (type II). The type of packing has a significant effect on the accessibility of the DNA, as evidenced by intercalation and light-harvesting experiments. Only the vesicles exhibiting the sheet-like DNA alignment are accessible for intercalation by ethidium bromide or for the integration of chromophore-labelled DNA via a strand exchange process. The dynamic nature of type I vesicles enables their elaboration into artificial light-harvesting complexes by DNA-guided introduction of Cy3-acceptor chromophores. DNA-constructed vesicles of the kind shown here represent versatile intermediates that are amenable to further modification for tailored nanotechnology applications.

The Watson-Crick complementarity of DNA strands that leads to their pairing into the DNA double helix, and that enables DNA to serve so effectively as genetic material, can be used for other purposes, too. This chapter describes the awesome skill of DNA to direct the assemblies of more complex DNA structural motifs than simple double, triple, or quadruple helical forms of DNA described in the introductory chapter of this book.

Cells of higher eukaryotes process within minutes double strand breaks (DSBs) in their genome using a non-homologous end joining (NHEJ) apparatus that engages DNA-PKcs, Ku, DNA ligase IV, XRCC4 and other as of yet unidentified factors. Although chemical inhibition, or mutation, in any of these factors delays processing, cells ultimately remove the majority of DNA DSBs using an alternative pathway operating with an order of magnitude slower kinetics. This alternative pathway is active in mutants deficient in genes of the RAD52 epistasis group and frequently joins incorrect ends. We proposed, therefore, that it reflects an alternative form of NHEJ that operates as a backup (B-NHEJ) to the DNA-PK-dependent (D-NHEJ) pathway, rather than homology directed repair of DSBs. The present study investigates the role of Ku in the coordination of these pathways using as a model end joining of restriction endonuclease linearized plasmid DNA in whole cell extracts. Efficient, error-free, end joining observed in such in vitro reactions is strongly inhibited by anti-Ku antibodies. The inhibition requires DNA-PKcs, despite the fact that Ku efficiently binds DNA ends in the presence of antibodies, or in the absence of DNA-PKcs. Strong inhibition of DNA end joining is also mediated by wortmannin, an inhibitor of DNA-PKcs, in the presence but not in the absence of Ku, and this inhibition can be rescued by pre-incubating the reaction with double stranded oligonucleotides. The results are compatible with a role of Ku in directing end joining to a DNA-PK dependent pathway, mediated by efficient end binding and productive interactions with DNA-PKcs. On the other hand, efficient end joining is observed in extracts of cells lacking DNA-PKcs, as well as in Ku-depleted extracts in line with the operation of alternative pathways. Extracts depleted of Ku and DNA-PKcs rejoin blunt ends, as well as homologous ends with 3' or 5' protruding single strands with similar efficiency, but addition of Ku suppresses joining of blunt ends and homologous ends with 3' overhangs. We propose that the affinity of Ku for DNA ends, particularly when cooperating with DNA-PKcs, suppresses B-NHEJ by quickly and efficiently binding DNA ends and directing them to D-NHEJ for rapid joining. A chromatin-based model of DNA DSB rejoining accommodating biochemical and genetic results is presented and deviations between in vitro and in vivo results discussed.

The robust ability of DNA to self-assemble into miscellaneous nanostructures, as it was described in the previous chapter, can also be employed for the construction of molecular size active nanomechanical devices and nanorobots, aka molecular machines or nanomachines, made entirely or partially of DNA. This idea is so inspiring that in the past decade there was a burst of activity in the area of DNA machines: a diverse variety of them have been designed based on several different principles, and their workability has been proved using different physical techniques.

We provided an ultrasensitive sensing strategy for microRNA detection by first employing branched DNA. With the aid of microcontact printing, we realized the multiplex sensing of different kinds of liver cancer biomarkers: microRNA and protein simultaneously. Delicately designed branched DNA included multiple complementary sticky ends as probe to microRNA capture and the double-stranded rigid branched core to increase the active sticky-ends distance and expose more DNA probes for sensitivity. The branched DNA enables 2 orders of magnitude increase in sensitivity for microRNA detection over single-stranded DNA. The limit of detection reaches as low as 10 attomolar (S/N = 3) for miR-223 and 10–12 M for α-fetoprotein. In addition, this system shows high selectivity and appropriate reproducibility (the relative standard deviation is less than 20%) in physiological media. Serum samples are tested and the results of α-fetoprotein are in good agreement with the current gold-standard method, electrochemiluminesce...

Liquid crystals (LCs) ordering of DNA and RNA oligomers relies on the presence of inter-duplex end-to-end attraction, driving the formation of linear aggregates. Such interactions are gauged, at a macroscopic level, by the osmotic pressure at the isotropic-nematic and nematic-columnar phase transitions. We studied aqueous solutions of PEG and DNA duplex-forming oligomers, finding that there is a wide range of concentrations in which these mixtures phase separate into coexisting PEG-rich and DNA-rich phases, the latter being either in the isotropic state or ordered as a nematic or columnar LC. We determined the phase diagram in mixtures of PEG and DNA duplexes with different terminal motifs – blunt ends, sticky overhangs, aggregation-preventing overhangs – and measured the partitioning of the species in the coexisting phases. On this basis, we determined the osmotic pressure as a function of the DNA concentration across the phase diagram. We compared the equation of state obtained in this way with ...

DNA double-strand breaks (DSBs) pose an everyday threat to the conservation of genetic information and therefore life itself. Several pathways have evolved to repair these cytotoxic lesions by rejoining broken ends, among them the nonhomologous end-joining mechanism that utilizes a DNA ligase. Here, we use a custom-designed DNA origami nanostructure as a model system to specifically mimic a DNA DSB, enabling us to study the end-joining of two fluorescently labeled DNA with the T4 DNA ligase on the single-molecule level. The ligation reaction is monitored by Forster resonance energy transfer (FRET) experiments both in solution and on surface-anchored origamis. Due to the modularity of DNA nanotechnology, DNA double strands with different complementary overhang lengths can be studied using the same DNA origami design. We show that the T4 DNA ligase repairs sticky ends more efficiently than blunt ends and that the ligation efficiency is influenced by both DNA sequence and the incubation conditions. Before ligation, dynamic fluctuations of the FRET signal are observed due to transient binding of the sticky overhangs. After ligation, the FRET signal becomes static. Thus, we can directly monitor the ligation reaction through the transition from dynamic to static FRET signals. Finally, we revert the ligation process using a restriction enzyme digestion and religate the resulting blunt ends. The here-presented DNA origami platform is thus suited to study complex multistep reactions occurring over several cycles of enzymatic treatment.

The encapsulation of folded proteins in stabilizing matrices is one of the challenges of soft‐matter materials science. Capturing such fragile bio‐macromolecules from aqueous solution, and embedding them in a lattice that stabilizes them against denaturation and decomposition is difficult. Here, we report that tetrahedral oligonucleotide hybrids as branching elements, and connecting DNA duplexes with sticky ends can assemble into materials. The material‐forming property was used to capture DNA‐binding proteins selectively from aqueous protein mixtures. The three‐dimensional networks also encapsulate guest molecules in a size‐selective manner, accommodating proteins up to a molecular weight of approximately 159 kDa for the connecting duplex lengths tested. Exploratory experiments with green fluorescent protein showed that, when embedded in the DNA‐based matrix, the protein is more stable toward denaturation than in the free form, and retains its luminescent properties for at least 90 days in dry form. The noncrystalline biohybrid matrices presented herein may be used for capturing other proteins or for producing functional materials.

A new colorimetric DNAzyme nanowire sensor has been developed for the sensitive and selective quantification of lipopolysaccharides (LPS). The sensor was a label-free and isotheral reaction system based on a LPS-binding aptamer and autonomously assembled DNAzyme nanowire. In the sensing system, an aptamer-initiator (AI) was designed to recognize the LPS and initiated the hybridization chain reaction (HCR) of two DNA hairpins (H1 and H2). All the H2 containing sticky ends formed terminal deoxynucleotidyl transferase (TdT) −induced G-quadruplexes. In the presence of LPS, on end of AI, and LPS-aptamer, formed the LPS/aptamer complex with the remaining LPS, the other end of AI, the initiator then started the alternate-opening of H1 and H2 through HCR. Furthermore, TdT-induced hemin/G-quadruplex DNAzymes along DNA nanowires were self-assembled after HCR process. As a result, the solution in light yellow was observed via the addition of H2O2/TMB. After optimization, the time to detect the signal was approximately 40 min, and the reaction temperature was in a broad range from 4 °C to 37 °C. The detection platform showed excellent sensitivity and selectivity for LPS from E. coli O111:B4 in a series of samples. In less than 2 h, a colorimetric response was achieved and the concentrations of LPS detected could be as low as 100 pg/mL. The semi-quantitative observation limit was 20 ng/mL with the naked eye. A drinking water sample was tested, which further demonstrated the feasibility of the proposed method applied for biological samples.

It is known that DNA molecules can be used to build a various of complicated geometrical DNA nanostructures with programmable sequence design, and these DNA nanomaterials show a promising application in biotechnology and biomedicine. However, the construction of large-sized three dimensional DNA-based nanomaterials still remains a challenge. In this work, we propose a new strategy that only employs one target DNA to trigger multiple catalyzed hairpin assembly (CHA) reactions and sticky ends self-assembly to prepare hundreds of nanometer-sized DNA nanoparticles. Moreover, the obtained DNA nanoparticles can be served as efficient biosensors for sensitive colorimetric nucleic acids detection with a detection limit of 7.7 pM

Human telomerase is an endogenous ribonucleoprotein that is over-expressed in most types of malignant cancer cells. Sensitive and specific detection of telomerase activity is crucial for better understanding its role in cancer cells and further exploring its function in cancer diagnosis. Here, we develop convertible DNA ends-based silver nanoprobes for sensitive and specific colorimetric detection telomerase activity. Silver nanoprobes are constructed by modifying telomerase binding substrates (TS) that are pre-hybridized with complementary sequences onto silver nanoparticles (AgNPs), via the coordination between consecutive cytosines in TS strand and AgNPs. This forms blunt-end terminated, double-stranded DNA on the surface of AgNPs. Under the action of telomerase, TS on the silver nanoprobes are elongated with telomeric repeats, converting DNA stiff blunt ends to flexible single-stranded dangling ends. The dangling ends enhance the stability of nanoprobes and relieve their salt-induced aggregation, and the solution shows a yellow color. When telomerase is inactive, the blunt end-terminated nanoprobes cannot resist salt-induced aggregation, resulting in a gray color of solution. Based on telomerase-regulated DNA “blunt-dangling” ends conversion-induced AgNPs' dispersity and color change, colorimetric detection of the endogenous telomerase with AgNPs is realized. The detection limit is equivalent to 1 cell/μL of telomerase activity, and extracts from cancer cells and normal cells are visually distinguished through color difference. The proposed strategy will offer a new approach for reliable, convenient quantification of telomerase activity in biochemical research and clinical diagnosis.

The authors describe a strategy for colorimetric detection of DNA. It is making use of a target catalyzed DNA nanostructure assembly and gold nanoparticles. The assay comprises the following steps: (a) A programmed DNA nanostructure is assembled from three auxiliary hairpin structure DNAs and the single stranded DNA (ssDNA; the target/analyte); (b) in the presence of target DNA, these three hairpin DNAs are opened, thereby activating a catalytic self-assembly process via a target assisted toe-hold strand displacement reaction; (c) The formed DNA nanostructures are mixed with gold nanoparticles. As the DNA nanostructure is less stabilized without ssDNA sticky ends, it cannot prevent the gold nanoparticles (AuNPs) to undergo salt-induced aggregation which is accompanied by a color change from purple to blue; (d) The color change of the colloid solution can be read out with bare eyes or instrumentally. The detection limit by using UV–vis spectrometry is 0.6 pM of target DNA. This is comparable to previously AuNP-based methods. Thus, this assay provides free modification detection based on DNA nanostructure without sophisticated procedures. Conceivably, the method can be applied to numerous other DNA targets.

Long-range ordering of DNA crossover tiles with blunt ends on lipid bilayers is investigated using atomic force microscopy. "Blunt-ended" tiles do not have single-stranded complementary ends, and thus instead of assembling via base-pairing, they can interact by π-stacking of their duplex ends. This work demonstrates that the balance of base π-stacking interactions between the ends of DNA duplexes, cholesterol-mediated DNA anchoring, and electrostatic DNA binding to supported lipid bilayers (SLBs) presents an opportunity to build dynamic materials with long-range order on a soft support. The tiles are shown to organize into novel tunable surface packing morphologies on the micrometer scale. This work focuses on three-point star (3PS) tiles that are either unmodified or modified with a cholesterol unit and investigates their interactions on supported lipid bilayers. On fluid bilayers, the cholesterol tiles form extended hexagonal arrays with few defects, while the unmodified tiles do not bind. In contrast, both modified and unmodified tiles bind to gel-phase bilayers and produce arrays of new organized morphologies. With increasing tile concentration, we observe a range of motifs, that progressively favor tile-tile packing over duplex-end π-stacking. These structures can selectively pattern domains of phase-separated lipid bilayers, and the patterning is also observed for four-arm cross-tiles. Dynamic blunt end contacts promote error correction and network reconfiguration to maximize favorable interactions with the substrate and are required for the observed tile organization. These results suggest that small blunt-ended tiles can be used as a platform to organize oligonucleotides, nanoparticles, and proteins into extensive networks at the interface with biologically relevant membrane systems or other soft surface materials for applications in cellular recognition, plasmonics, light harvesting, model systems for membrane protein assemblies, or analytical devices.

Cell-sized liposomes and droplets coated with lipid layers have been used as platforms for understanding live cells, constructing artificial cells, and implementing functional biomedical tools such as biosensing platforms and drug delivery systems. However, these systems are very fragile, which results from the absence of cytoskeletons in these systems. Here, we construct an artificial cytoskeleton using DNA nanostructures. The designed DNA oligomers form a Y-shaped nanostructure and connect to each other with their complementary sticky ends to form networks. To undercoat lipid membranes with this DNA network, we used cationic lipids that attract negatively charged DNA. By encapsulating the DNA into the droplets, we successfully created a DNA shell underneath the membrane. The DNA shells increased interfacial tension, elastic modulus, and shear modulus of the droplet surface, consequently stabilizing the lipid droplets. Such drastic changes in stability were detected only when the DNA shell was in the gel phase. Furthermore, we demonstrate that liposomes with the DNA gel shell are substantially tolerant against outer osmotic shock. These results clearly show the DNA gel shell is a stabilizer of the lipid membrane akin to the cytoskeleton in live cells.

The generalized structure of DNA recombination is based on the rules of rough Set. Inthis paper we are discussing the relationship between rough Set and DNA recombination. Weconstruct a new recombination operator using the properties of DNA and RNA. Using the processof cutting and sticking of a sequence of genes, new types of topological structures are constructedand some of their properties and characterization are investigated. We studied recombinationoperators in the statement” The restriction enzyme cuts both molecules at the same sequence("Sticky" End)”.

A two-way colorimetric biosensor based on unmodified gold nanoparticles (GNPs) and a switchable double-stranded DNA (dsDNA) concatemer have been demonstrated. Two hairpin probes (H1 and H2) were first designed that provided the fuels to assemble the dsDNA concatemers via hybridization chain reaction (HCR). A functional hairpin (FH) was rationally designed to recognize the target sequences. All the hairpins contained a single-stranded DNA (ssDNA) loop and sticky end to prevent GNPs from salt-induced aggregation. In the presence of target sequence, the capture probe blocked in the FH recognizes the target to form a duplex DNA, which causes the release of the initiator probe by FH conformational change. This process then starts the alternate-opening of H1 and H2 through HCR, and dsDNA concatemers grow from the target sequence. As a result, unmodified GNPs undergo salt-induced aggregation because the formed dsDNA concatemers are stiffer and provide less stabilization. A light purple-to-blue color variation was observed in the bulk solution, termed the light-off sensing way. Furthermore, H1 ingeniously inserted an aptamer sequence to generate dsDNA concatemers with multiple small molecule binding sites. In the presence of small molecule targets, concatemers can be disassembled into mixtures with ssDNA sticky ends. A blue-to-purple reverse color variation was observed due to the regeneration of the ssDNA, termed the light-on way. The two-way biosensor can detect both nucleic acids and small molecule targets with one sensing device. This switchable sensing element is label-free, enzyme-free, and sophisticated-instrumentation-free. The detection limits of both targets were below nanomolar.

Multifunctional viral protein (VP35) encoded by the highly pathogenic Ebola viruses (EBOVs) can antagonize host double-stranded RNA (dsRNA) sensors and immune response because of the simultaneous recognition of dsRNA backbone and blunt ends. Mutation of select hydrophobic conserved basic residues within the VP35 inhibitory domain (IID) abrogates its dsRNA-binding activity, and impairs VP35-mediated interferon (IFN) antagonism. Herein the detailed binding mechanism between dsRNA and WT, single mutant, and double mutant were investigated by all-atom molecular dynamics (MD) simulation and binding energy calculation. R312A/R322A double mutations results in a completely different binding site and orientation upon the structure analyses. The calculated binding free energy results reveal that R312A, R322A, and K339A single mutations decrease the binding free energies by 17.82, 13.18, and 13.68 kcal mol-1 , respectively. The binding energy decomposition indicates that the strong binding affinity of the key residues is mainly due to the contributions of electrostatic interactions in the gas phase, where come from the positively charged side chain and the negatively charged dsRNA backbone. R312A, R322A, and K339A single mutations have no significant effect on VP35 IID conformation, but the mutations influence the contributions of electrostatic interactions in the gas phase. The calculated results reveal that end-cap residues which mainly contribute VDW interactions can recognize and capture dsRNA blunt ends, and the central basic residues (R312, R322, and K339) which mainly contribute favorable electrostatic interactions with dsRNA backbone can fix dsRNA binding site and orientation. Proteins 2017; 85:1008-1023. © 2017 Wiley Periodicals, Inc.

In the present work, an enzyme-free and signal amplified nucleic acids detection method based on the fluorescent nucleotide base analog pyrrolo-deoxycytidine (P-dC) and catalyzed hairpin assembly (CHA) has been developed. In the CHA signal amplification system, two hairpin auxiliary probes, H1 and H2, which containing a fluorescent P-dC at the end of the stem, respectively, were used as the fluorescent probes. The fluorescence of P-dC in H1 and H2 was quenched owing to the stacking interaction among the bases in the stem. In the presence of the target DNA, the catalytic assembly of H1 and H2 was triggered and the target could be released during the helix DNA H1-H2 complex formation process, then the released target was used to trigger another reaction cycle. In the H1-H2 complex, P-dC was located in the flexible single-stranded DNA (ssDNA) sticky ends instead of the rigid stem, thus resulting in the increase of fluorescence. The cycling use of the target in the CHA system amplified the fluorescence signal, and the detection limit of this method was obtained as 19pM, which is 3 orders of magnitude sensitive than the conventional fluorescent nucleotide base analogs based approach without CHA signal amplification.

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. Moreover, the length of rigidity core can also be tuned to result in different dihedral angles between the component helices of a certain motif therefore different numbers of component helices. The extended set of motifs is used for self-assembly of complex one dimensional, two dimensional and three dimensional structures.

Tile-based self-assembly is a robust system in the construction of three-dimensional DNA nanostructures but it has been rarely applied to other helical biopolymers. β-Glucan is an immunoactive natural polymer which exists in a triple helical conformation. Herein, we report that β-glucan, after modification using two types of short chain acyl groups, can self-assemble into tiles with inactivated sticky ends at the interface of two solvents. These tiles consist of a single layer of helices laterally aligned, and the sticky ends can be activated when a few acyl groups at the ends are removed; these tiles can further pack into mesoporous nanocapsules, in a similar process as the sticky DNA tiles pack into complex polyhedral nano-objects. These nanocapsules were found to have targeted effects to antigen presenting cells in a RAW264.7 cell model. Our study suggests that tile-based self-assembly can be a general strategy for helical biopolymers, and on fully exploiting this strategy, various new functional nanostructures will become accessible in the future.