Parallel, double-helix DNA nanostructures using interstrand cross-linked oligonucleotides with bismaleimide linkers.
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Development of a nanostructured DNA delivery scaffold via electrospinning of PLGA and PLA-PEG block copolymers.
The present work utilizes electrospinning to fabricate synthetic polymer/DNA composite scaffolds for therapeutic application in gene delivery for tissue engineering. The scaffolds are non-woven, nano-fibered, membranous structures composed predominantly of poly(lactide-co-glycolide) (PLGA) random copolymer and a poly(D,L-lactide)-poly(ethylene glycol) (PLA-PEG) block copolymer. Release of plasmid DNA from the scaffolds was sustained over a 20-day study period, with maximum release occurring at approximately 2 h. Cumulative release profiles indicated amounts released were approximately 68-80% of the initially loaded DNA. Variations in the PLGA to PLA-PEG block copolymer ratio vastly affected the overall structural morphology, as well as both the rate and efficiency of DNA release. Results indicated that DNA released directly from these electrospun scaffolds was indeed intact, capable of cellular transfection, and successfully encoded the protein beta-galactosidase. When tested under tensile loads, the electrospun polymer/DNA composite scaffolds exhibited tensile moduli of approximately 35 MPa, with approximately 45% strain initially. These values approximate those of skin and cartilage. Taken together, this work represents the first successful demonstration of plasmid DNA incorporation into a polymer scaffold using electrospinning.
Animals↗
DNA-programmed assembly of nanostructures.
DNA is a unique material for nanotechnology since it is possible to use base sequences to encode instructions for assembly in a predetermined fashion at the nanometre scale. Synthetic oligonucleotides are readily obtained by automated synthesis and numerous techniques have been developed for conjugating DNA with other materials. The exact spatial positioning of materials is crucial for the future development of complex nanodevices and the emerging field of DNA-nanotechnology is now exploring DNA-programmed processes for the assembly of organic compounds, biomolecules, and inorganic materials.
DNA↗
Hybridization with nanostructures of single-stranded DNA.
Nanostructures of single-stranded DNA (ssDNA) were produced within alkanethiol self-assembled monolayers using nanografting, an atomic force microscopy (AFM) based lithography technique. Next, variations of the fabrication parameters, such as the concentration of ssDNA or lines per frame, allowed for the regulation of the density of ssDNA molecules within the nanostructures. The label-free hybridization of nanostructures, monitored using high-resolution AFM imaging, has proven to be highly selective and sensitive; as few as 50 molecules can be detected. The efficiency of the hybridization reaction at the nanometer scale highly depends on the ssDNA packing density within the nanostructures. This investigation provides a fundamental step toward sensitive DNA detection and construction of complex DNA architectures on surfaces.
DNA, Single-Stranded↗
Discrete nanostructures of quantum dots/Au with DNA.
Nanostructures of colloidal CdSe/ZnS core/shell quantum dots (QDs) surrounded by a discrete number of Au nanoparticles were generated via DNA hybridization and purified by gel electrophoresis. Statistics from TEM analysis showed a high yield of designed structures. The distance between Au particles and QD, the number of Au around the central QD, and the size of Au and QD can be adjusted. Rationally designed structures such as these hold great promise for researching the physical interactions between semiconductor and Au nanoparticles and for developing more efficient nanoprobes.
DNA↗
Fabrication of Au-DNA-Au nanostructure with new-type DNA-Au conjugate.
Nano devices fabricated from DNA and nanoparticles found many important applications in both biotechnology and nanotechnology. Conventionally, DNA was linked to gold surfaces through a linker. In this study, DNAs were directly attached to gold surfaces through S-Au bonds by modifying 5'-end of DNA with a mercapto group. The pH of the reaction mixture played an important role in DNA-gold conjugate fabrication. The DNA-Au conjugates were successfully obtained in pure forms and, with the use of this new type conjugate, Au-DNA-Au nanostructure was prepared. The Au-DNA-Au nanostructure was characterized by AFM.
DNA↗
Atomic force microscopy study of the interaction of DNA and nanostructured beta-Gallia rutile.
The ability to attach DNA molecules to solid planar substrates is desired for imaging the molecule and for building DNA-mediated nanostructures. The deposition of DNA on [001] rutile and beta-gallia rutile (BGR) substrates from buffer solutions containing various divalent cations was studied using tapping mode atomic force microscopy (AFM). beta-Gallia rutile intergrowths were prepared by spin-coating gallium isopropoxide onto [001]-oriented TiO2 single-crystal slabs and heating above 1350 degrees C for >24 h, resulting in the formation of intergrowth lines along the {210} planes in the parent rutile structure. Rutile and BGR intergrowth substrates were exposed to various buffered solutions containing DNA and the following divalent cations: Ca(II), Co(II), Cu(II), Fe(II), Mg(II), Mn(II), Ni(II), and Zn(II). Among all the cations examined, only Ni(II) resulted in the attachment of DNA on the rutile surfaces. DNA attachment to BGR surfaces was strong enough to allow AFM imaging when the deposition buffer contained one of the following cations: Co(II), Mg(II), Mn(II), Ni(II), and Zn(II). For all of these cations, DNA attachment occurred preferentially, but not exclusively, along BGR intergrowth lines. When buffers without cation additions and those containing Ca(II), Cu(II), and Fe(II) were used, DNA failed to bind the BGR surfaces strongly enough to allow AFM imaging. The mechanism(s) by which DNA attaches to the BGR surface is (are) not well understood but may involve the incorporation of divalent cations at the tunnel sites of the BGR intergrowths.
DNA↗
DNA assembly using bis-peptide nucleic acids (bisPNAs).
DNA nanostructures are ordered oligonucleotide arrangements that have applications for DNA computers, crystallography, diagnostics and material sciences. Peptide nucleic acid (PNA) is a DNA/RNA mimic that offers many advantages for hybridization, but its potential for application in the field of DNA nanotechnology has yet to be thoroughly examined. We report the synthesis and characterization of tethered PNA molecules (bisPNAs) designed to assemble two individual DNA molecules through Watson-Crick base pairing. The spacer regions linking the PNAs were varied in length and contained amino acids with different electrostatic properties. We observed that bisPNAs effectively assembled oligonucleotides that were either the exact length of the PNA or that contained overhanging regions that projected outwards. In contrast, DNA assembly was much less efficient if the oligonucleotides contained overhanging regions that projected inwards. Surprisingly, the length of the spacer region between the PNA sequences did not greatly affect the efficiency of DNA assembly. Reasons for inefficient assembly of inward projecting DNA oligonucleotides include non-sequence-specific intramolecular interactions between the overhanging region of the bisPNA and steric conflicts that complicate simultaneous binding of two inward projecting strands. These results suggest that bisPNA molecules can be used for self-assembling DNA nanostructures provided that the arrangement of the hybridizing DNA oligonucleotides does not interfere with simultaneous hybridization to the bisPNA molecule.
Chromatography, High Pressure Liquid↗
Self-assembly of DNA-streptavidin nanostructures and their use as reagents in immuno-PCR.
The self-assembly of bis-biotinylated double-stranded DNA and the tetravalent biotin-binding protein streptavidin (STV) have been studied by non-denaturing gel electrophoresis and atomic force microscopy (AFM). The rapid self-assembly reproducibly generated populations of individual oligomeric complexes. Most strikingly, the oligomers predominantly contained bivalent STV molecules bridging two adjacent DNA fragments to form linear nanostructures. Trivalent STV branch points occurred with a lower frequency and the presence of tetravalent STV was scarce. However, valency distribution, size and the exchange dynamics of the supramolecular aggregates were highly sensitive to stoichiometric variations in the relative molar coupling ratio of bis-biotinylated DNA and STV. The largest aggregates were obtained from equimolar amounts while excess STV led to the formation of smaller oligomers appearing as fingerprint-like band patterns in electrophoresis. Excess DNA, however, induces a complete breakdown of the oligomers, likely a consequence of the instability of STV conjugates containing more than two biotinylated DNA fragments. It was demonstrated that the oligomers can further be functionalized, for instance by the coupling of biotinylated immunoglobulins. Both pure and also antibody-modified DNA-STV oligomers were used as reagents in immuno-PCR (IPCR), a highly sensitive detection method for proteins and other antigens. Employment of the supramolecular reagents led to an approximately 100-fold enhanced sensitivity compared to the conventional IPCR procedure.
Base Sequence↗
Nanostructured electrochemical DNA biosensors for detection of the effect of berberine on DNA from cancer cells.
Multi walled carbon nanotubes (MWNT) in dimethylformamide (DMF) or aqueous sodium dodecyl sulfate (SDS) solution, colloidal gold nanoparticles (GNP) in phosphate buffer solution (PBS), and a GNP-MWNT mixture in aqueous SDS solution have been investigated for chemical modification of a screen-printed carbon electrode used as the signal transducer of a dsDNA-based biosensor. Differential pulse voltammetry of the DNA redox marker Co[(phen)3]3+ and the guanine moiety anodic oxidation and cyclic voltammetry with K3[Fe(CN)6] as indicator revealed substantial enhancement of the response of the biosensor, particularly when MWNT in SDS solution was used. The biosensor was used in testing of berberine, an isoquinoline plant alkaloid with significant antimicrobial and anticancer activity. Berberine had a very strong, concentration-dependent, effect on the structural stability of DNA from the human cancer cells (U937 cells) whereas non-cancer cells were changed only when berberine concentrations were relatively high 75 and 50 microg mL(-1).
Berberine↗
DNA nanomachines and nanostructures involving quadruplexes.
DNA is an attractive component for molecular recognition, because of its self-assembly properties. Its three-dimensional structure can differ markedly from the classical double helix. For example, DNA or RNA strands carrying guanine or cytosine stretches associate into four-stranded structures called G-quadruplexes or i-DNA, respectively. Since 2002, several groups have described nanomachines that take advantage of this structural polymorphism. We first introduce the unusual structures that are involved in these devices (i.e., i-DNA and G-quadruplexes) and then describe the opening and closing steps that allow cycling. A quadruplex-duplex molecular machine is then presented in detail, together with the rules that govern its formation, its opening/closing kinetics and the various technical and physico-chemical parameters that play a role in the efficiency of this device. Finally, we review the few examples of nanostructures that involve quadruplexes.
Base Sequence↗
Charge transfer through DNA nanoscaled assembly programmable with DNA building blocks.
DNA nanostructures based on programmable DNA molecular recognition have been developed, but the nanoelectronics of using DNA is still challenging. A more rapid charge-transfer (CT) process through the DNA nanoassembly is required for further development of programmable DNA nanoelectronics. In this article, we present direct absorption measurements of the long-range CT over a 140-A DNA assembly based on a GC repetitive sequence constructed by simply mixing DNA building blocks. We show that a CT through DNA nanoscale assembly is possible and programmable with the designed DNA sequence.
Base Sequence↗
Deposition of DNA rafts on cationic SAMs on silicon [100].
We demonstrate a guided self-assembly approach to the fabrication of DNA nanostructures on silicon substrates. DNA oligonucleotides self-assemble into "rafts" 8 x 37 x 2 nm in size. The rafts bind to cationic SAMs on silicon wafers. Electron-beam lithography of a thin poly(methyl methacrylate) (PMMA) resist layer was used to define trenches, and (3-aminopropyl)triethoxysilane (APTES), a cationic SAM precursor, was deposited from aqueous solution onto the exposed silicon dioxide at the trench bottoms. The remaining PMMA can be cleanly stripped off with dichloromethane, leaving APTES layers 0.7-1.2 nm in thickness and 110 nm in width. DNA rafts bind selectively to the resulting APTES stripes. The coverage of DNA rafts on adjacent areas of silicon dioxide is 20 times lower than on the APTES stripes. The topographic features of the rafts, measured by AFM, are identical to those of rafts deposited on wide-area SAMs. Binding to the APTES stripes appears to be very strong as indicated by "jamming" of the rafts at a saturation coverage of 42% and the stability to repeated AFM scanning in air.
Microscopy, Atomic Force↗
Nanostructured dna-protein aggregates consisting of covalent oligonucleotide-streptavidin conjugates.
Covalent conjugates consisting of streptavidin and a 24-mer single-stranded DNA oligonucleotide have been oligomerized by cross-linking with a 5',5'-bis-biotinylated 169-base-pair double-stranded DNA (dsDNA) fragment. The oligomeric conjugates formed have been analyzed by nondenaturing gel electrophoresis and scanning-force microscopy (SFM). The comparison of analogous oligomers, prepared from native STV and the bis-biotinylated dsDNA fragment, revealed that the covalent STV-oligonucleotide hybrid conjugates self-assemble to generate oligomeric aggregates of significant smaller size, containing on average only about 2.5 times less dsDNA fragments per aggregate. Likely, this is a consequence of electrostatic or steric repulsion between the dsDNA and the single-stranded oligomer covalently attached to the hybrid, as indicated from control experiments. Nevertheless, the single-stranded oligonucleotide moiety within the oligomeric conjugates can be used as a selective molecular handle for further functionalization and manipulation. For instance, it was used for specific DNA-directed immobilization at a surface, previously functionalized with complementary capture oligonucleotides. Moreover, we demonstrate that macromolecules, such as STV and antibody molecules, which are tagged with the complementary oligonucleotide, specifically bind to the supramolecular DNA-STV oligomeric conjugates. This leads to a novel class of functional DNA-protein conjugates, suitable, for instance, as reagents in immuno-PCR or as building blocks in molecular nanotechnology.
Animals↗
DNA-based machines.
Nucleic acids include substantial information in their base sequence and their hybridization-complexation motifs. Recent research efforts attempt to utilize this biomolecular information to develop DNA nanostructures exhibiting machine-like functions. DNA nano-assemblies revealing tweezers, motor, and walker activities exemplify a few such machines. The DNA-based machines provide new components that act as sensitive sensors, transporters, or drug delivery systems.
Base Sequence↗
DNA tile based self-assembly: building complex nanoarchitectures.
DNA tile based self-assembly provides an attractive route to create nanoarchitectures of programmable patterns. It also offers excellent scaffolds for directed self-assembly of nanometer-scale materials, ranging from nanoparticles to proteins, with potential applications in constructing nanoelectronic/nanophotonic devices and protein/ligand nanoarrays. This Review first summarizes the currently available DNA tile toolboxes and further emphasizes recent developments toward self-assembling DNA nanostructures with increasing complexity. Exciting progress using DNA tiles for directed self-assembly of other nanometer scale components is also discussed.
Base Composition↗
Transition of nanostructure in DNA-cationic surfactant complexes with the added salt.
Nanostructures of complexes of DNA with single-chain surfactant of octadecyltrimethylammonium (OTA) and double-chain surfactant of didodecyldimethylammonium (DDA) in aqueous NaCl solution at concentration, Cs, from 0 to 500 mM were studied using small-angle-scattering techniques (SAXS). SAXS profiles of the DNA-OTA complex show two SAXS peaks with a spacing ratio of 1:3(1/2) in the solution at Cs below 150 mM and three peaks with a spacing ratio of 1:3(1/2):4(1/2) at Cs above 250 mM. Contents of Na+ and Cl- ions in the complexes evaluated from the atomic absorbance for Na+ and the potentiometry for Cl- revealed charge molar ratios of OTA/DNA = 1 and DDA/DNA = 1.25. Contents of Na+ and Cl- ions per ionic unit of DNA molecule in the DNA-OTA complex equilibrating with the solution at Cs below 100 mM were much less than 0.1, while they increased with NaCl concentration at Cs above 200 mM. The DNA-OTA complex in the solution at Cs above 260 mM exhibited an endothermic peak in the DSC measurements, and the others did not. On the basis of the experimental results, the salt concentration dependent nanostructures are discussed.
Cations↗
Self-assembly of symmetric finite-size DNA nanoarrays.
We report a novel and cost-effective strategy to self-assemble finite-size DNA nanoarrays. This strategy takes advantage of the geometric symmetry of the DNA nanostructures. In general, to construct a 2D array with a total of N tiles containing Cm symmetry, where m = 2, 3, 4, or 6, the number of unique tiles the fixed-size array requires is N/m, if N/m is an integral number, or Int(N/m) + 1, if N/m is an nonintegral number. We herein demonstrate two examples of fixed-size arrays with C2 and C4-fold symmetry.dagger
DNA↗