Supplies
Staple strands for the DNA origami seeds have been bought from Sigma-Aldrich (desalt purification). SST strands for the SST sublattices have been bought from Sangon Biotech (PAGE purification). The sequences of all strands could be present in Supplementary Tables 1–6. Scaffold DNA strands (p7560, CS3-L and CS4) have been bought from Tilibit Nanosystems. All DNA strands have been saved at −20 °C after being dissolved in ultrapure water. Different chemical substances have been bought from Sigma-Aldrich. Carbon-coated TEM grids have been bought from Ted Pella. Quantifoil grids have been bought from Quantifoil Micro Instruments GmbH. AFM suggestions have been bought from Bruker.
Preparation of DNA origami seeds
DNA origami seeds have been designed utilizing caDNAno software program. Seed S is a DNA origami bundle consisting of 48 helices (8-helix × 8-helix rim, 4-helix × 4-helix pore) organized in a sq. array (Fig. 1b(i)). Seed H is a DNA origami bundle composed of 54 helices (12-helix × 6-helix rim, 6-helix × 3-helix pore) organized in a honeycomb array (Fig. 1b(ii)). The hole seed design facilitates the correct identification of the seed inside a DNA moiré superlattice for microscopy characterizations. The twisted geometries of the DNA origami seeds and the twist angles of the Z2 (or Z4) segments have been simulated utilizing the SNUPI program28. The simulation parameters are offered in Supplementary Fig. 3. Place vectors on the seeds have been calculated on the premise of coordinates extracted from the SNUPI simulation output information. To assemble the DNA origami seeds, staple strands have been blended with scaffold strands in a molar ratio of 10:1 in TE-MgCl2 buffer (10 mM Tris, 1 mM EDTA and 20 mM MgCl2, pH 7.8). The combination was then annealed in a PCR thermocycler utilizing the next protocol: 65 °C for 20 min; from 60 °C to 40 °C at 40 min per diploma Celsius; and from 40 °C to 25 °C at 15 min per diploma Celsius.
Agarose gel evaluation and pattern purification
The DNA origami seeds have been electrophoresed on a 1.0 % agarose gel containing 1× GelRed in 0.5× TBE buffer with 11 mM Mg2+ for 3 h at 80 V with ice cooling. The goal band was excised from the gel and squeezed between two glass slides. The focus of the purified DNA origami seeds was decided utilizing ultraviolet–seen absorption spectroscopy. All purified DNA origami seeds have been saved at 4 °C earlier than additional use.
Development of unseeded DNA sublattice monolayers
The sq. and honeycomb lattices are created utilizing 32-nt SSTs with 4 8-nt domains and 36-nt SSTs with 4 9-nt domains, respectively. The kagome lattice is fashioned utilizing two 32-nt SST variants: one adopts a U-shape, as within the sq. lattice, whereas the opposite is linear, comprising three domains (8 nt, 16 nt and eight nt). To kind the lattice monolayers, the SST strands have been blended in an equimolar stoichiometric ratio from a 200 µM inventory in TE buffer (10 mM Tris and 1 mM EDTA, pH 7.8) supplemented with 40 mM MgCl2. The combination was then annealed in a PCR thermocycler utilizing the next protocol: 65 °C for 20 min; 48 °C to 47 °C at 12 h per diploma Celsius for the honeycomb lattice; 44 °C to 43 °C at 12 h per diploma Celsius for the kagome lattice; and 39 °C to 38 °C at 12 h per diploma Celsius for the sq. lattice.
Development of seeded monolayers, bilayers and trilayers
The SST strands (1.2 µM), purified DNA origami seeds (0.1–1.1 nM) and MgCl2 (remaining focus of 40 mM) have been individually preheated for five min at correct temperatures (honeycomb, 50 °C; kagome, 48 °C; sq., 45 °C). They have been then blended shortly to keep away from temperature fluctuations and incubated for five min. Lastly, seeded development proceeded on the designated nucleation and development temperatures.
Dynamic mild scattering measurements of homogeneous nucleation kinetics
Dynamic mild scattering experiments have been performed utilizing Zetasizer Professional (Malvern Devices). The SST strands have been pipetted right into a cuvette, and the dimensions of particular person SST strands was measured. To acquire their nucleation kinetics, the SST strands and 40 mM MgCl2 have been preheated for five min after which blended. Measurements have been taken instantly on the combination at 15-s intervals.
Optical microscopy imaging
A 1 cm × 1 cm silicon wafer was cleaned sequentially with Milli-Q water, acetone and ethanol. It was then dried utilizing nitrogen gasoline. Afterwards, the wafer was handled with plasma cleansing (30 mA, 5 min) to make sure a hydrophilic floor. Tenfold diluted DNA sublattices or superlattices have been absorbed for 1 h onto the freshly ready silicon wafer. Then,75% (vol/vol), 90% (vol/vol) and 100% (vol/vol) ethanol was sequentially used to clean the floor after absorption. Lastly, the wafer was dried utilizing nitrogen gasoline.
SEM imaging
Tenfold diluted DNA sublattices or superlattices have been absorbed for 1 h onto the freshly ready 1 cm × 1 cm silicon wafer. The wafer was then stained for 30 s utilizing a 2% aqueous uranyl formate resolution containing 25 mM NaOH. Then, 75% (vol/vol), 90% (vol/vol) and 100% (vol/vol) ethanol was sequentially used to clean the floor after absorption. Lastly, the wafer was dried utilizing nitrogen gasoline. SEM imaging was carried out utilizing Raith eLine Plus.
AFM imaging
The silicon wafers with stained DNA samples have been straight used for AFM imaging with a ScanAsyst-air tip in air mode.
TEM imaging
5 microlitres of purified DNA origami seeds (1 nM) or tenfold diluted DNA sublattices or superlattices have been absorbed for 10 min onto a glow discharged, carbon-coated TEM grid. The grids have been then stained for 10 s utilizing a 2% aqueous uranyl formate resolution containing 25 mM NaOH. TEM imaging was carried out utilizing Phillip CM 200 TEM operated at 200 kV. STEM photographs have been carried out utilizing JEOL JEM-ARM200F operated at 60 kV, outfitted with a chilly field-emission gun and a probe Cs corrector (DCOR, CEOS GmbH).
Cryo-EM characterization
Three microlitres of DNA lattices have been pipetted onto glow-discharged Quantifoil grids. The pattern was plunge-frozen utilizing a Vitrobot Mark IV at 20 °C and humidity of 90%, with a wait time of 0 s, blot time of 6 s, blot pressure of −1 and drain time of 0 s. Imaging was carried out utilizing Talos Arctica (Thermo Fisher Scientific) operated at 200 kV, outfitted with a Falcon III detector. Class-average cryo-EM photographs have been obtained utilizing the EMAN2 software program bundle, model 2.99.47.
All-atom molecular dynamics simulations
Molecular dynamics simulations have been carried out utilizing the Gromacs2021 software program bundle. The AMBER99sb pressure discipline was used to explain the interactions throughout the system. The simulation field was constructed utilizing Gromacs, with all elements of the system being correctly positioned in response to the necessities. The field was then crammed with water, using the TIP3P water mannequin. Mg2+ and Cl− have been added to neutralize the system, making certain that it was total cost impartial. Earlier than the manufacturing run, the system underwent vitality minimization adopted by a 100-ps equilibration simulation to permit correct rest. The manufacturing molecular dynamics simulation used the leapfrog algorithm to combine Newton’s equations of movement, with an integration time step of 0.002 ps and a complete of fifty,000,000 steps, leading to a complete simulation time of 100 ns. The V-rescale thermostat was used to keep up the simulation temperature at 318.15 Okay, and the Parrinello–Rahman barostat was utilized to maintain the strain fixed at 1.0 bar. The Verlet scheme was used for neighbour looking out. The cut-off radius for Coulombic interactions was set to 1.2 nm, with the long-range electrostatic interactions being corrected utilizing the particle mesh Ewald technique.
Statistics and reproducibility
No statistical technique was used to predetermine pattern dimension. No knowledge have been excluded from the analyses. The experiments weren’t randomized. The investigators weren’t blinded to allocation throughout experiments and end result evaluation.
