Available via license: CC BY-NC-ND 3.0
Content may be subject to copyright.
A preview of the PDF is not available
DNA nanotechnology
Abstract and Figures
Since Watson and Crick’s determination of its structure nearly 50 years ago, DNA has come to fill our lives in many areas, from genetic counseling to forensics, from genomics to gene therapy. These, and other ways in which DNA affects human activities, are related to its function as genetic material, not just our genetic material, but the genetic material of all living organisms. Here, we will ignore DNA’s biological role; rather, we will discuss how the properties that make it so successful in acting as genetic material also make it a convenient and logical molecule to use for constructing new materials on the nanometer scale. The well-known B-DNA double helix is about 20 Å wide, and its helical repeat is 10-10.6 nucleotide pairs for a pitch of 34–36 Å. Thus, constructions made from DNA will have nanoscale features.
We are all aware that the DNA found in cells is a double helix consisting of two antiparallel strands held together by specific hydrogen-bonded base pairs; adenine (A) always pairs with thymine (T), and guanine (G) always pairs with cytosine (C). The specificity of this base pairing and the ability to ensure that it occurs in this fashion (and not some other1) is key to the use of DNA in materials applications. The double helical arrangement of the two molecules leads to a linear helix axis, linear not in the geometrical sense of being a straight line, but in the topological sense of being unbranched. Genetic engineers discovered in the 1970s how to splice together pieces of DNA to add new genes to DNA molecules2, and synthetic chemists worked out convenient syntheses for short pieces of DNA (up to ∼100–150 units) in the 1980s3. Regardless of the impact of these technologies on biological systems, hooking together linear molecules leads only to longer linear molecules, with circles, knots, and catenanes perhaps resulting from time to time.
Figures - available via license: Creative Commons Attribution-NonCommercial-NoDerivs 3.0 Unported
Content may be subject to copyright.
... Since its invention two decades ago [4], toeholdmediated DNA strand displacement has been a central mechanism for programming dynamical function in DNA nanotechnology [5,6]. As shown in figure 1A, a stable complex of two strands can be reconfigured such that an invading strand replaces the original partner via a branch migration process. ...
Tile displacement is a newly-recognized mechanism in DNA nanotechnology that exploits principles analogous to toehold-mediated strand displacement but within the context of self-assembled DNA origami tile arrays. Here, we formulate an abstract model of tile displacement for the simplest case: individual assemblies interacting with monomer tiles in solution. We give several constructions for programmable computation by tile displacement, from circuits to cellular automata, that vary in how they use energy (or not) to drive the system forward (or not), how much space and how many tile types they require, and whether their computational power is limited to PTIME or PSPACE with respect to the size of the system. In particular, we show that tile displacement systems are Turing universal and can simulate arbitrary two-dimensional synchronous block cellular automata, where each transition rule for updating the state of a 2 by 2 neighborhood is implemented by just a single tile.
DNA self-assembly and in particular DNA origami has evolved into a reliable workhorse for organizing organic and inorganic materials with nanometer precision and with exactly controlled stoichiometry. To ensure the intended performance of a given DNA structure it is beneficial to determine its folding temperature, which in turn yields the best possible assembly of all DNA strands. Here we show that temperature-controlled sample holders and standard fluorescence spectrometers or dynamic light scattering (DLS) set-ups in a static light scattering (SLS) configuration allow for monitoring the assembly progress in real time. With this robust label-free technique we determine the folding and melting temperatures of a set of different DNA origami structures without the need for more tedious protocols. In addition, we use the method to follow digestion of DNA structures in the presence of DNase I and find strikingly different resistances towards enzymatic degradation depending on the structural design of the DNA object.
DNA with data encoding and molecular recognition is rarely used in combination with electrochemistry for multipurpose integrated applications (especially in sensing, information communication and security). Herein, we demonstrated an electrochemical aptasensing, information communication and safety system for detection of fish pathogens (Aeromonas hydrophila or Edwardsiella tarda) and molecular information encryption and hiding. Two fish pathogens can be easily and quickly detected by electrochemistry, respectively, with high selectivity and sensitivity (detection limit lower than 1 cfu/mL) without the need for traditional time-consuming biochemical culturing process. The specific interaction of the probe (DNA aptamer) with targets (pathogens) on the tiny and imperceptible electrochemical platform provides protection for hiding DNA aptamers containing the encoded message, but also offers a foundation for developing of molecular cryptography and steganography. This electrochemical system, which is similar to mail communication, does not record information on paper, but a molecular mail that records information through DNA and reads information using electrochemical sensing, or more precisely, molecular electrochemical mail (namely molecular ‘email’). Our study proved that the combination of the recognition and encoding capabilities of DNA aptamers with electrochemistry can open a new door for molecular-level digitization technology. In the future, large-capacity, easy-to-operate, resettable, and flexible molecular crypto-steganography will be developed for molecular cascade communication and control.
The systematic analysis and precise manipulation of a variety of biomolecules should lead to unprecedented findings in fundamental biology. However, conventional technology cannot meet the current requirements. Despite this, there has been progress as DNA nanotechnology has evolved to generate DNA nanostructures and circuits over the past four decades. Many potential applications of DNA nanotechnology for live cell measurements have begun to emerge owing to the biocompatibility, nanometer addressability, and stimulus responsiveness of DNA. In this review, the DNA nanotechnology‐empowered live cell measurements which are currently available are summarized. The stability of the DNA nanostructures, in a cellular microenvironment, which is crucial for accomplishing precise live cell measurements, is first summarized. Thereafter, measurements in the extracellular and intracellular microenvironment, in live cells, are introduced. Finally, the challenges that are innate to, and the further developments that are possible in this nascent field are discussed. Cell measurements play important roles in biology. Recently, the applications of DNA nanotechnology for live cell measurements have begun to emerge. In this review, DNA nanotechnology‐empowered measurements for live cells are summarized. The review is expected to inspire future works about live cell measurements based on DNA nanotechnology.
Seeman's pioneer idea has led to the foundation of DNA nanostructures, resulting in a remarkable advancement in DNA nanotechnology. Over the last few decades, remarkable advances in drug delivery techniques have resulted in the self-assembly of DNA molecules for encapsulating candidate drug molecules. The beauty of DNA nanostructures is their nuclear targeting capability with high spatial addressability and tremendous potential for active nucleus targeting. However, effectively programming and assembling those DNA molecules remains a challenge, making the path to DNA nanostructures for real-world applications difficult. Because of their small size, most nanostructures are self-capable of infiltrating into the tumor cellular environment. Furthermore, to enable controlled and site-specific delivery of encapsulated drug molecules, DNA nanostructures are functionalized with special moieties that allow them to bind specific targets and release cargo at only targeted sites rather than non-specific sites, resulting in the prevention/limitation of cellular toxicity. In light of this, the current review seeks to shed light on the versatility of the DNA molecule as a targeting and encapsulating moiety for active drug molecules in order to achieve controlled and specific drug release with spatial and temporal precision. Furthermore, this review focused on the challenges associated with the construction of DNA nanostructures as well as the most recent advances in DNA nanostructures functionalization using various materials for controlled and targeted delivery of medications for cancer therapy.
Self-assembly of biomolecules is ubiquitous in nature, which provides structural and functional machinery for the cells. During the past decades, material science has been greatly inspired by the assembly principles in nature to create artificial structures, opening new directions for biomedical applications. In this chapter, the self-assembly of deoxyribonucleic acid (DNA) polymer is shown to the readers as a novel way for nanofabrication that brings the encoded DNA polymers to aggregation into desired structures, which are increasingly used for biomedical applications for their diverse structures and functions. Specifically, the usage of self-assembled DNA structures at the bio-interfaces will be focused on, including the extracellular, cell plasma, and intracellular applications. Based on various structures and functions of these DNA assemblies, potential implications for diagnosis, tissue engineering, biosensing and imaging, drug delivery, and mechanism studies are presented. Therefore, this chapter will highlight the self-assembled DNA structures at the bio-interfaces, from principles to applications, in a way that may help future investigators, researchers, students, and stakeholders who intend to perform their research with DNA assemblies for biomedical applications.
This article covers the major themes in the pursuits of DNA-assisted nanoparticle assembly, with a special emphasis on methodological developments and their underlying driving motives and basic principles. Following a brief introduction, the topics are organized by starting from technical advancements in DNA-conjugated nanoparticle building blocks, continuing with the construction of static and dynamic nanostructures ranging from discrete nanoparticle oligomers to periodic crystals, and ending with some synergistic functions realized by DNA-programmed nanoassemblies and corresponding applications in various aspects. An outlook is finally given on existing challenges and future opportunities of this research area.
The kinetics of DNA hybridization are fundamental to biological processes and DNA-based technologies. However, the precise physical mechanisms that determine why different DNA sequences hybridize at different rates are not well understood. Secondary structure is one predictable factor that influences hybridization rates but is not sufficient on its own to fully explain the observed sequence-dependent variance. In this context, we measured hybridization rates of 43 different DNA sequences that are not predicted to form secondary structure and present a parsimonious physically justified model to quantify our observations. Accounting only for the combinatorics of complementary nucleating interactions and their sequence-dependent stability, the model achieves good correlation with experiment with only two free parameters. Our results indicate that greater repetition of Watson–Crick pairs increases the number of initial states able to proceed to full hybridization, with the stability of those pairings dictating the likelihood of such progression, thus providing new insight into the physical factors underpinning DNA hybridization rates.
The base pairs in double helical nucleic acids have been compared to see how they can be recognized by proteins. We conclude that a single hydrogen bond is inadequate for uniquely identifying any particular base pair, as this leads to numerous degeneracies. However, using two hydrogen bonds, fidelity of base pair recognition may be achieved. We propose specific amino-acid side chain interactions involving two hydrogen bonds as a component of the recognition system for base pairs. In the major groove we suggest that asparagine or glutamine binds to adenine of the base pair or arginine binds to guanine. In the minor groove, we suggest an interaction between asparagine or glutamine with guanine of the base pair. We also discuss the role that ions and other amino-acid side chains may play in recognition interactions.
A polyamide nucleic acid (PNA) was designed by detaching the deoxyribose phosphate backbone of DNA in a computer model and
replacing it with an achiral polyamide backbone. On the basis of this model, oligomers consisting of thymine-linked aminoethylglycyl
units were prepared. These oligomers recognize their complementary target in double-stranded DNA by strand displacement. The
displacement is made possible by the extraordinarily high stability of the PNA-DNA hybrids. The results show that the backbone
of DNA can be replaced by a polyamide, with the resulting oligomer retaining base-specific hybridization.
The crystal structure of d-CGACGATCGT has been determined to a resolution of 2.6 Å. The molecule was synthesized by standard phosphoramidite procedures, and purified by anion-exchange HPLC. Crystals are monolclinic, space group P21, with unit cell dimensions, a = 26.45 Å, b = 34.66 Å, c = 32.17 Å, β = 113.45° and Z = 4, containing a B-DNA double helix in each crystallographic asymmetric unit. The structure was solved using molecular replacement, aided by an isomorphous derivative, in which a bromine atom was attached to the 5 position of cytosine 8. Problems of fit between the search model and the structure ultimately obtained necessitated the use of Patterson correlation procedures between the determination of the orientation and the translation of the molecule. In all, 69 solvent molecules have been identified, and the structure has been refined to an R-factor of 0.214, using the 1421 reflections with F>2σ(F), collected at −120°C. The sequence produces a molecule containing eight Watson-Crick base-pairs and a two-nucleotide 5′-sticky end at each end of the duplex. The sticky ends cohere with one another, so the molecules form continuous 10-fold double helices throughout the crystal, with each strand being interrupted by inherent staggered nicks. The relative angular relationships between helices in the structure differ from each other; most of the arrangements differ from Holliday junctions, whose rotational orientations are phased by a crossover and which are modeled to contain double helices that are exactly parallel or antiparallel. However, one helical juxtaposition in this crystal is similar to the alignment of double helices in parallel Holliday junctions. A survey of DNA decamers that also form infinite helices in crystals reveals relationships that approximate both parallel and antiparallel Holliday junction alignments.
A two-dimensional DNA crystal has been designed and constructed from Holliday junction analogues that contain two helical domains twisted relative to each other. The Holliday junction is not an inherently rigid system, but it can be made less flexible if it is combined into a larger construct. We have fused four junctions into a rhombus-like molecule consisting of four six-turn helices, two on an upper layer and two on a lower layer; the branch points, which define vertices, are separated by four double helical turns each. Ligation of the rhombus-like motifs produces no cyclic species, when assayed by ligation-closure experiments. Self-assembly of the rhombuses in one dimension leads to a linear pattern. The rhombuses can be directed to self-assemble by hydrogen bonding into a two-dimensional periodic array, whose spacing is six turns in each direction. The expected spacing is seen when the array is observed by atomic force microscopy (AFM). Variation of the dimensions of the repeat unit from six turns × six turns to six turns × eight turns results in the expected increase in unit cell dimensions. Hence, it is possible to assemble periodic arrays with tunable cavities using these components. This system also provides the opportunity to measure directly the angles or torsion angles between the arms of branched junctions; here we measure the torsion angle between the helical domains of the Holliday junction analogue. We find by AFM that the torsion angle between helices is 63.5°, in good agreement with previous estimates.
A covalently closed molecular complex whose double-helical edges have the connectivity of a truncated octahedron has been assembled from DNA on a solid support. This three-connected Archimedean solid contains six squares and eight hexagons, formed from 36 edges arranged about 24 vertices. The vertices are the branch points of four-arm DNA junctions, so each vertex has an extra exocyclic arm associated with it. The construct contains six single-stranded cyclic DNA molecules that form the squares and the extra arms; in addition, there are eight cyclic strands that correspond to the eight hexagons. The molecule contains 1440 nucleotides in the edges and 1110 in the extra arms; the estimated molecular weight for the 2550 nucleotides in the construct is 790 kDa. Each edge contains two turns of double-helical DNA, so that the 14 strands form a catenated structure in which each strand is linked twice to its neighbors along each edge. Synthesis is proved by demonstrating the presence of each square in the object and then by confirming that the squares are flanked by tetracatenane substructures, corresponding to the hexagons. The success of this synthesis indicates that this technology has reached the stage where the control of topology is in hand, in the sense of both helix axis connectivity and strand linkage.
It is possible to generate sequences of oligomeric nucleic acids which will preferentially associate to form migrationally immobile junctions, rather than linear duplexes, as they usually do. These structures are predicated on the maximization of Watson-Crick base pairing and the lack of sequence symmetry customarily found in their analogs in living systems. Criteria are presented which oligonucleotide sequences must fulfill in order to yield these junction structures. The generable junctions are nexi, from which 3 to 8 double helices may emanate. Each junction may be treated as a macromolecular “valence cluster”, and the individual clusters may be linked together directly, or with pieces of linear DNA interspersed between them. This covalent linkage can be done with enormous specificity, using the sticky-ended ligation techniques currently employed in genetic engineering studies. It appears to be possible to generate covalently joined three-dimensional networks of nucleic acids which are periodic in connectivity and perhaps in space.
This paper extends the study and prototyping of unusual DNA motifs, unknown in nature, but founded on principles derived from biological structures. Artificially designed DNA complexes show promise as building blocks for the construction of useful nanoscale structures, devices, and computers. The DNA triple crossover (TX) complex described here extends the set of experimentally characterized building blocks. It consists of four oligonucleotides hybridized to form three double-stranded DNA helices lying in a plane and linked by strand exchange at four immobile crossover points. The topology selected for this TX molecule allows for the presence of reporter strands along the molecular diagonal that can be used to relate the inputs and outputs of DNA-based computation. Nucleotide sequence design for the synthetic strands was assisted by the application of algorithms that minimize possible alternative base-pairing structures. Synthetic oligonucleotides were purified, stoichiometric mixtures were annealed by slow cooling, and the resulting DNA structures were analyzed by nondenaturing gel electrophoresis and heat-induced unfolding. Ferguson analysis and hydroxyl radical autofootprinting provide strong evidence for the assembly of the strands to the target TX structure. Ligation of reporter strands has been demonstrated with this motif, as well as the self-assembly of hydrogen-bonded two-dimensional crystals in two different arrangements. Future applications of TX units include the construction of larger structures from multiple TX units, and DNA-based computation. In addition to the presence of reporter strands, potential advantages of TX units over other DNA structures include space for gaps in molecular arrays, larger spatial displacements in nanodevices, and the incorporation of well-structured out-of-plane components in two-dimensional arrays.
Recent work has demonstrated the self-assembly of designed periodic
two-dimensional arrays composed of DNA tiles, in which the
intermolecular contacts are directed by `sticky' ends. In a mathematical
context, aperiodic mosaics may be formed by the self-assembly of `Wang'
tiles, a process that emulates the operation of a Turing machine.
Macroscopic self-assembly has been used to perform computations; there
is also a logical equivalence between DNA sticky ends and Wang tile
edges. This suggests that the self-assembly of DNA-based tiles could be
used to perform DNA-based computation. Algorithmic aperiodic
self-assembly requires greater fidelity than periodic self-assembly,
because correct tiles must compete with partially correct tiles. Here we
report a one-dimensional algorithmic self-assembly of DNA
triple-crossover molecules that can be used to execute four steps of a
logical (cumulative XOR) operation on a string of binary bits.
Stable DNA branched junction molecules can be used as the building blocks for stick-figures in which the edges are double-helical DNA and the vertices correspond to the branch points of the junctions. Sticky-ended cohesion is used to direct the association of individual branched complexes. The sequences of these molecules are assigned by a sequence-symmetry minimization procedure. Successful ligation experiments include the oligomerization of individual three-arm and four-arm junctions, the assembly of a quadrilateral from four junctions with different sticky ends, and the recent construction of a molecule with the connectivity of a cube. Possible applications include the assembly of molecular electronic devices, the formation of macromolecular-scale zeolites to host biological complexes for diffraction analysis, and the development of new catalysts.
A principal goal of biotechnology is the assembly of novel biomaterials for analytical, industrial and therapeutic purposes. The advent of stable immobile nucleic acid branched junctions makes DNA a good candidate for building frameworks to which proteins or other functional molecules can be attached and thereby juxtaposed. The addition of single-stranded 'sticky' ends to branched DNA molecules converts them into macromolecular valence clusters that can be ligated together. The edges of these frameworks are double-helical DNA, and the vertices correspond to the branch points of junctions. Here, we report the construction from DNA of a covalently closed cube-like molecular complex containing twelve equal-length double-helical edges arranged about eight vertices. Each of the six 'faces' of the object is a single-stranded cyclic molecule, doubly catenated to four neighbouring strands, and each vertex is connected by an edge to three others. Each edge contains a unique restriction site for analytical purposes. This is the first construction of a closed polyhedral object from DNA.