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Submicrometre geometrically encoded fluorescent barcodes self-assembled from DNA

Nature Chemistry volume 4, pages 832–839 (2012)Cite this article

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Abstract

The identification and differentiation of a large number of distinct molecular species with high temporal and spatial resolution is a major challenge in biomedical science. Fluorescence microscopy is a powerful tool, but its multiplexing ability is limited by the number of spectrally distinguishable fluorophores. Here, we used (deoxy)ribonucleic acid (DNA)-origami technology to construct submicrometre nanorods that act as fluorescent barcodes. We demonstrate that spatial control over the positioning of fluorophores on the surface of a stiff DNA nanorod can produce 216 distinct barcodes that can be decoded unambiguously using epifluorescence or total internal reflection fluorescence microscopy. Barcodes with higher spatial information density were demonstrated via the construction of super-resolution barcodes with features spaced by ∼40 nm. One species of the barcodes was used to tag yeast surface receptors, which suggests their potential applications as in situ imaging probes for diverse biomolecular and cellular entities in their native environments.

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Figure 1: Design of a barcode based on a DNA nanorod.
Figure 2: Fluorescent barcodes with single-labelled zones.
Figure 3: Fluorescent barcodes with dual-labelled zones.
Figure 4: Super-resolution fluorescent barcodes.
Figure 5: Fluorescent barcode with nonlinear geometry.
Figure 6: Tagging yeast cells with the GRG barcodes as in situ imaging probes.

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Acknowledgements

We thank C. Steinhauer and S. P. Laurien for help with super-resolution microscopy software development, the Harvard Center for Biological Imaging, as well as the Nikon Imaging Center at Harvard Medical School, for the use of their microscopes and S. M. Douglas for providing the transmission electron microscopy images used in Supplementary Fig. S5. This work is supported by a National Institutes of Health (NIH) Director's New Innovator Award (1DP2OD007292), a National Science Foundation Faculty Early Career Development Award (CCF1054898), an Office of Naval Research Young Investigator Program Award (N000141110914), an Office of Naval Research grant (N000141010827) and a Wyss Institute for Biologically Engineering Faculty Startup Fund to P.Y., and an NIH Director's New Innovator Award (1DP2OD004641) and a Wyss Institute for Biologically Inspired Engineering Faculty Award to W.M.S. C. Li, D.L. and G.M.C. acknowledge support from the National Human Genome Research Institute, Centers of Excellence in Genomic Science. R.J. acknowledges support from the Alexander von Humboldt-Foundation through a Feodor Lynen fellowship.

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Author notes

  1. Andrew M. Leifer

    Present address: Present address: Lewis Sigler Institute for Integrative Genomics, Princeton University, Princeton, New Jersey 08544, USA,

Authors and Affiliations

  1. Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, 02115, Massachusetts, USA

    Chenxiang Lin, Ralf Jungmann, Chao Li, Daniel Levner, George M. Church, William M. Shih & Peng Yin

  2. Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, 02115, Massachusetts, USA

    Chenxiang Lin & William M. Shih

  3. Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, 02115, Massachusetts, USA

    Chenxiang Lin & William M. Shih

  4. Department of Systems Biology, Harvard Medical School, Boston, 02115, Massachusetts, USA

    Ralf Jungmann & Peng Yin

  5. Program in Biophysics, Harvard University, Cambridge, 02138, Massachusetts, USA

    Andrew M. Leifer

  6. Department of Genetics, Harvard Medical School, Boston, 02115, Massachusetts, USA

    Chao Li, Daniel Levner & George M. Church

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Contributions

C. Lin conceived the project, designed and conducted the majority of the experiments, analysed the data and prepared the majority of the manuscript. R.J. conceived the super-resolution barcode study, designed and conducted experiments for this study, analysed the data and prepared the manuscript. A.M.L. wrote the MATLAB script for the automated barcode deciphering and prepared the manuscript. C. Li wrote the C script for the barcode geometry characterization. D.L. (with C. Lin) performed the yeast-tagging experiment. G.M.C. championed multiplexed in situ, supervised C. Li and D.L., and critiqued the data and the manuscript. W.M.S. conceived the project, discussed the results and prepared the manuscript. P.Y. conceived, designed and supervised the study, interpreted the data and prepared the manuscript. All authors reviewed and approved the manuscript.

Corresponding authors

Correspondence to William M. Shih or Peng Yin.

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The authors declare no competing financial interests.

Supplementary information

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Supplementary information (PDF 20281 kb)

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High-resolution Fig. S3 (PDF 8483 kb)

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High-resolution Fig. S8 (PDF 11430 kb)

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High-resolution Fig. S9 (PDF 5958 kb)

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High-resolution Fig. S12 (PDF 5111 kb)

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Lin, C., Jungmann, R., Leifer, A. et al. Submicrometre geometrically encoded fluorescent barcodes self-assembled from DNA. Nature Chem 4, 832–839 (2012). https://doi.org/10.1038/nchem.1451

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