| Naturally,DNA is the heritable material to store genetic information with the specificity of the Watson-Crick base pairing interaction.Nadrian C.Seeman,in the 1980 s,proposed and executed the use of DNA as a molecular building block to construct geometrically defined nanostructures.Later on,in 2006,Rothemund developed a method for construction of DNA nanostructures called DNA origami,in which a virus-based single-stranded(ss)DNA(M13mp18 genomic DNA)was folded into desired shapes and patterns along with short synthetic staple strands.Development of DNA nanotechnology has been driven by applications to achieve complex structures of high yield but with simple manufacturing process.A wide range of different DNA nanostructures and machines in one,two and three-dimensions(1D,2D,and 3D)have been developed,as well as DNA nanostructures that incorporate other functional materials,such as proteins or metallic nanoparticles.Potential applications of DNA nanotechnology are wide,for example,patterning nano-electronics,immobilising proteins for specific recognition,exploring biomacromolecule interactions,and even mimicking logical devices for computing.Mostly often,people use the whole set of liner DNA strands for construction of 1D,2D and 3D nanostructures,whereas our group use circularised DNAs as central scaffold strands for assembling DNA nanostructures.Recently we applied circularised DNA dumbbell molecules as the central scaffold strand for construction of different types of tiles and different kinds of nanostructures,which include ribbons,lattices,and nanotubes.Details are described as follows:1.DNA dumbbell tiles with uneven widths for 2D arrays: In the first project,DNA dumbbell molecules were synthesised using a conventional method: a pair of two hairpin oligonucleotides associate with each other by sticky end cohesion to form a dumbbell structure,and then the two nicks located at the central duplex stem are covalently connected with T4 ligase.Two differently-sized(A of 86-nt(nucleotides)and B of 96-nt)dumbbell oligonucleotides were designed and synthesised.The former has an 11 bp long central double-stranded(ds)stem and two ss 32-nt head loops;the latter has a 16 bp long central ds stem and two ss 32-nt head loops.After the synthesis and purification of DNA dumbbells,we used them as scaffold strands to synthesize DNA dumbbell tiles and 2D arrays with the help of additional linear strands,so-called helper strands.We only used a one-tile system and one-pot or two-step protocols to demonstrate the self-assembly approach for dumbbell tiles and 2D arrays.Totally four tiles(A_O,A_E,B_O & B_E)were prepared by construction of two from each dumbbell and four 2D arrays of array-A_O,array-A_E,array-B_O,and array-B_E,were grown into narrow nanoribbons and nanotubes with the widest width up to 200 nm.Because the randomly sequenced central stem is only half-width of the head motifs,its mechanic strength is also half of the head motifs.Therefore,the assembled 2D arrays are metastable and only stay for a week in the buffer solution.2.2D DNA lattice arrays assembled from DNA dumbbell tiles using poly(A-T)-rich stems: In the second project,because the poly(A-T)-rich duplexes have been confirmed to be more rigid than randomly-sequenced ones under acidic environments,to strengthen the rigidity of the central stem,we synthesised four DNA dumbbell molecules by using the new poly(A-T)-rich sequences as stems,namely,dumbbell molecules of D6,D11,D16,and D21,with 6,11,16,and 21 bp stems respectively.For specific association,the two head loops of a dumbbell molecule were designed to have different random sequences of ss 32 nt,while for comparison of 2D arrays,the two head loop sequences were copied for all the four DNA dumbbell molecules.We synthesised D11,D16,and D21 with the conventional approach,in which two hairpin molecules were associated with the stem together,then the two nicks formed by complementary sticky ends at the stem were ligated covalently by T4 ligase.DNA dumbbell molecule D6 was designed as a whole dumbbell molecule with only one nick at the stem for ligation because the 6 bp stem is too short to hold two nicks.DNA dumbbell molecules were used as scaffolds to construct dumbbell tiles after their synthesis and purification.Four DNA dumbbell molecules of D6,D11,D16,and D21 were used for construction of eight dumbbell tiles,namely,D6_O,D6_E,D11_O,D11_E,D16_O,D16_E,D21_O,and D21_E,where the subscripts of O and E represent the intertitle distance of odd and even half-turns in a 2D DNA array.To grow 2D lattices,we adapted the standard intertitle distance of 4(even number)and 5(odd number)half-turns for tile connection by sticky end cohesion and applied a one-tile system and a one-pot annealing approach.Totally eight 2D arrays were assembled,namely,array-D6_O,array-D6_E,array-D11_O,array-D11_E,array-D16_O,array-D16_E,array-D21_O,and array-D21_E.DNA 2D arrays were designed by assembling tiles into lattice structures.Array-D21_O and array-D21_E were observed to be mixtures of nanoribbons and nanotubes,all other arrays including array-D11_E were assembled as planar nanoribbons.The enhanced mechanical strength of central stems expands the choice of more stem lengths to grow DNA dumbbell tiles and stable,single-crystalline 2D arrays.The rigid stems,dumbbell tiles,and 2D arrays enable the high-resolution Atomic Force Microscopy(AFM)imaging of individual tiles at a lateral resolution of around 3 nm feasible,and thus the measurement of 2D Bravais lattice parameters of each array possible.All the measured lattice parameters are in line with their theoretical designs. |
