Cost-efficient folding of functionalized DNA origami nanostructures via staple recycling

DNA origami nanostructures are powerful molecular tools for the controlled arrangement of functional molecules and thus have important applications in biomedicine, sensing, and materials science. The fabrication of DNA origami nanostructures commonly requires a high excess of staple strands, leading to material waste and high costs, especially when large numbers of modified staples are to be incorporated. Here, we present a method for recycling non-modified as well as biotinylated and fluorophore-modified excess staple strands using molecular weight cut-off (MWCO) ultrafiltration and reusing them in subsequent folding reactions. The structural integrity of the folded DNA origami nanostructures as well as the incorporation and functionality of the introduced modifications are maintained over at least five folding cycles. The resulting reduction in staple costs due to staple recycling reaches 33% over five folding cycles, with a theoretical maximum of 41% for large numbers of cycles. This cost-effective and sustainable approach is straightforward to implement in any given DNA origami fabrication pipeline and particularly attractive for applications requiring large numbers of expensive modifications where substantial reductions in absolute costs can be achieved in this way.