Scaffolded DNA origami allows the programmable development of nanoscale buildings by means of the hybridization of a protracted single-stranded scaffold with tons of of quick staple strands. Nevertheless, the reliance on quite a few artificial oligonucleotides stays a key barrier to scalable and cost-effective manufacturing of DNA nanostructures. On this research, we introduce a long-staple design technique that extends the size of particular person staple strands to 100–200 nucleotides (nt), thereby decreasing the full variety of strands required whereas sustaining meeting effectivity and structural constancy. We show that this strategy is broadly suitable with a wide range of origami architectures, together with each manually designed lattice-based buildings and algorithmically generated wireframe geometries, with out requiring adjustments to well-established design workflows. Utilizing consultant 2D and 3D buildings, we present that lengthy staples can assemble effectively underneath the identical thermal annealing situations and Mg2+ concentrations as quick staples, yielding closing buildings with comparable morphology. To additional help organic manufacturing of staple strands, we generated lengthy staples by way of rolling circle amplification (RCA) utilizing custom-designed round templates, every encoding a particular lengthy staple sequence. This modular design permits for versatile and selective synthesis of desired staples, both individually or in pooled codecs. These RCA-derived staples have been efficiently utilized in construction meeting, confirming the feasibility of enzyme-based synthesis for long-staple designs. This modular and adaptable technique affords a sensible route towards scalable fabrication of practical DNA nanostructures throughout various design frameworks.
