Scaffolded DNA origami allows the programmable building of nanoscale constructions by the hybridization of an extended single-stranded scaffold with tons of of brief 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 examine, 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 display that this strategy is broadly suitable with a wide range of origami architectures, together with each manually designed lattice-based constructions and algorithmically generated wireframe geometries, with out requiring modifications to well-established design workflows. Utilizing consultant 2D and 3D constructions, we present that lengthy staples can assemble effectively beneath the identical thermal annealing situations and Mg2+ concentrations as brief staples, yielding last constructions with comparable morphology. To additional assist 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 had 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 numerous design frameworks.
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