Nick-and-Digest Strategy for Programmable Circular ssDNA Production and Scalable DNA Origami Assembly.

ABSTRACT: DNA origami enables the precise self-assembly of complex nanostructures with applications in drug delivery, biosensing, and nanoelectronics. However, the scalability of DNA origami is constrained by the limited length of the available single-stranded DNA (ssDNA) scaffolds. Here, we present a sequence-flexible "nick-and-digest" strategy to generate long circular ssDNA (cssDNA) scaffolds of customizable lengths directly from plasmid DNA. By combining Cas9 (D10A) nickase (Cas9n) with T7 exonuclease (T7 Exo), we generate high-purity cssDNA scaffolds of around 7,000 and 15,000 nucleotides (7k-nt and 15k-nt) with minimal sequence dependence. These extended scaffolds enable the one-pot folding of large-scale origami structures (147 × 107 nm) that double the surface area of conventional 7 kb desig

SUBMITTER: Ye J 

PROVIDER: S-EPMC12648328 | biostudies-literature | 2025 Nov

REPOSITORIES: biostudies-literature