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Multi-micron crisscross structures grown from DNA-origami slats

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An Author Correction to this article was published on 23 March 2023

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Abstract

Living systems achieve robust self-assembly across a wide range of length scales. In the synthetic realm, nanofabrication strategies such as DNA origami have enabled robust self-assembly of submicron-scale shapes from a multitude of single-stranded components. To achieve greater complexity, subsequent hierarchical joining of origami can be pursued. However, erroneous and missing linkages restrict the number of unique origami that can be practically combined into a single design. Here we extend crisscross polymerization, a strategy previously demonstrated with single-stranded components, to DNA-origami ‘slats’ for fabrication of custom multi-micron shapes with user-defined nanoscale surface patterning. Using a library of ~2,000 strands that are combinatorially arranged to create unique DNA-origami slats, we realize finite structures composed of >1,000 uniquely addressable slats, with a mass exceeding 5 GDa, lateral dimensions of roughly 2 µm and a multitude of periodic structures. Robust production of target crisscross structures is enabled through strict control over initiation, rapid growth and minimal premature termination, and highly orthogonal binding specificities. Thus crisscross growth provides a route for prototyping and scalable production of structures integrating thousands of unique components (that is, origami slats) that each is sophisticated and molecularly precise.

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Fig. 1: Overview of crisscross assembly of DNA-origami slats.
Fig. 2: Assembly of finite megastructures from DNA-origami slats, where every slat is unique and addressable (see the designs in Fig. 1c).
Fig. 3: Assembly of periodic ribbons and sheets that grow with 6HB slats in one and two dimensions.
Fig. 4: Finite and periodic crisscross megastructures as addressable DNA canvases to pattern arbitrary cargo.
Fig. 5: Characterization of growth versus reaction parameters.

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Data availability

All raw TEM image data that were measured to determine growth and nucleation of origami crisscross megastructures are available upon request from W.M.S.

Code availability

Scripts that were used to make various assignments of handles of staple oligonucleotides, and scripts that were used to measure Hamming distances of sequence assignments, are available at https://github.com/aersh/origamicrisscross.

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Acknowledgements

We would like to thank the following individuals: J. Kishi for suggesting and helping write a grant for the Echo Acoustic Liquid Handler that made this work possible; S. Cabi and T. Zhang for helping test early designs of crisscross origamis; and V. Manoharan, M. Brenner, R. Sørensen and J. Hahn for the fruitful discussions. Funding was provided by a Wyss Core Faculty Award (W.S., P.Y.); a Wyss Molecular Robotics Initiative Award (W.S., P.Y.); National Science Foundation DMREF Award 1435964 (W.S.); National Science Foundation Award CCF-1317291 (W.S.); Office of Naval Research Award N00014-15-1-0073 (W.S.); Office of Naval Research Award N00014-18-1-2566 (W.S.); Office of Naval Research DURIP Award N00014-19-1-2345 (W.S.); NIH NIGMS Award 5R01GM131401 (W.S.); NSERC PGSD3-502356-2017 (C.M.W.); Alexander S. Onassis Scholarship for Hellenes (A.E.).

Author information

Author notes

  1. Hiroshi M. Sasaki

    Present address: 10x Genomics, Inc., Pleasanton, CA, USA

  2. F. Eduardo Corea-Dilbert

    Present address: Geisel School of Medicine at Dartmouth, Hanover, NH, USA

  3. These authors contributed equally: Christopher M. Wintersinger, Dionis Minev, Anastasia Ershova.

Authors and Affiliations

  1. Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, MA, USA

    Christopher M. Wintersinger, Dionis Minev, Anastasia Ershova, Hiroshi M. Sasaki, Gokul Gowri, Jonathan F. Berengut, Peng Yin & William M. Shih

  2. John A. Paulson School of Engineering and Applied Sciences, Harvard University, Boston, MA, USA

    Christopher M. Wintersinger & Dionis Minev

  3. Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, MA, USA

    Christopher M. Wintersinger, Dionis Minev, Anastasia Ershova, Gokul Gowri, Jonathan F. Berengut, F. Eduardo Corea-Dilbert & William M. Shih

  4. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA, USA

    Christopher M. Wintersinger, Dionis Minev, Anastasia Ershova, Gokul Gowri & William M. Shih

  5. Department of Systems Biology, Harvard Medical School, Boston, MA, USA

    Hiroshi M. Sasaki, Gokul Gowri & Peng Yin

  6. EMBL Australia Node for Single Molecule Science, School of Medical Sciences, The University of New South Wales, Sydney, NSW, Australia

    Jonathan F. Berengut

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Contributions

Conceptualization: C.M.W., D.M., A.E., J.F.B., W.M.S. Methodology: C.M.W., D.M., A.E., W.M.S. Software: C.M.W., D.M., A.E., G.G. Validation: C.M.W., D.M., A.E. Formal analysis: C.M.W., D.M., A.E., H.M.S., G.G. Investigation: C.M.W., D.M., A.E., H.M.S., G.G., J.F.B., F.E.C.D. Writing (original draft): C.M.W. Writing (review and editing): C.M.W., D.M., A.E., H.M.S., G.G., F.E.C.D., W.M.S. Visualization: C.M.W., D.M., A.E., H.M.S., J.F.B. Supervision: C.M.W., P.Y., W.M.S. Funding acquisition: C.M.W., D.M., A.E., P.Y., W.M.S.

Corresponding author

Correspondence to William M. Shih.

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Competing interests

A patent (PCT/US2017/045013) entitled ‘Crisscross Cooperative Self-assembly’ has been filed based on this work.

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Supplementary information

Supplementary Information

Supplementary Text 1–8, Figs. 1–45, Tables 1–8 and references.

Supplementary Table

DNA sequences in Excel format

Supplementary Video 1

Supplementary Video 1

Supplementary Data 5–8

Supplementary Data 5–8

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Wintersinger, C.M., Minev, D., Ershova, A. et al. Multi-micron crisscross structures grown from DNA-origami slats. Nat. Nanotechnol. 18, 281–289 (2023). https://doi.org/10.1038/s41565-022-01283-1

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