Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Brief Communication
  • Published:

Imaging individual mRNA molecules using multiple singly labeled probes

Nature Methods volume 5, pages 877–879 (2008)Cite this article

Abstract

We describe a method for imaging individual mRNA molecules in fixed cells by probing each mRNA species with 48 or more short, singly labeled oligonucleotide probes. This makes each mRNA molecule visible as a computationally identifiable fluorescent spot by fluorescence microscopy. We demonstrate simultaneous detection of three mRNA species in single cells and mRNA detection in yeast, nematodes, fruit fly wing discs, and mammalian cell lines and neurons.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on SpringerLink
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Simultaneous detection of a unique sequence and a repeated sequence in individual mRNA molecules.
Figure 2: Simultaneous imaging of three different mRNAs in mammalian cells.
Figure 3: Imaging localized mRNAs in C. elegans and D. melanogaster.

Similar content being viewed by others

References

  1. Kaufmann, B.B. & van Oudenaarden, A. Curr. Opin. Genet. Dev. 17, 107–112 (2007).

    Article  CAS  Google Scholar 

  2. St. Johnston, D. Nat. Rev. Mol. Cell Biol. 6, 363–375 (2005).

    Article  CAS  Google Scholar 

  3. Gall, J.G. Proc. Natl. Acad. Sci. USA 60, 553–560 (1968).

    Article  CAS  Google Scholar 

  4. Levsky, J.M. & Singer, R.H. J. Cell Sci. 116, 2833–2838 (2003).

    Article  CAS  Google Scholar 

  5. Raap, A.K. et al. Hum. Mol. Genet. 4, 529–534 (1995).

    Article  CAS  Google Scholar 

  6. Femino, A.M., Fay, F.S., Fogarty, K. & Singer, R.H. Science 280, 585–590 (1998).

    Article  CAS  Google Scholar 

  7. Maamar, H., Raj, A. & Dubnau, D. Science 317, 526–529 (2007).

    Article  CAS  Google Scholar 

  8. Femino, A.M., Fogarty, K., Lifshitz, L.M., Carrington, W. & Singer, R.H. Methods Enzymol. 361, 245–304 (2003).

    Article  CAS  Google Scholar 

  9. Randolph, J.B. & Waggoner, A.S. Nucleic Acids Res. 25, 2923–2929 (1997).

    Article  CAS  Google Scholar 

  10. Vargas, D.Y., Raj, A., Marras, S.A., Kramer, F.R. & Tyagi, S. Proc. Natl. Acad. Sci. USA 102, 17008–17013 (2005).

    Article  CAS  Google Scholar 

  11. Raj, A., Peskin, C.S., Tranchina, D., Vargas, D.Y. & Tyagi, S. PLoS Biol. 4, e309 (2006).

    Article  Google Scholar 

  12. Wang, J.C. et al. Proc. Natl. Acad. Sci. USA 101, 15603–15608 (2004).

    Article  CAS  Google Scholar 

  13. Fukushige, T., Hawkins, M.G. & McGhee, J.D. Dev. Biol. 198, 286–302 (1998).

    CAS  PubMed  Google Scholar 

  14. Sanicola, M., Sekelsky, J., Elson, S. & Gelbart, W.M. Genetics 139, 745–756 (1995).

    CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We thank S.A.E. Marras, F.R. Kramer, D. Vargas, S. Sinha, E. Andersen, G. Neuert and Q. Yang. This work was supported by the US National Institute of Mental Health grant MH079197; National Institute of General Medicine grants GM068957, GM077183 and GM070357; and National Science Foundation grant PHY0548484. A.R. is supported by a National Science Foundation fellowship DMS-0603392 and a Burroughs Wellcome Fund Career Award at the Scientific Interface.

Author information

Authors and Affiliations

  1. Department of Physics, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, 02139, Massachusetts, USA

    Arjun Raj, Scott A Rifkin & Alexander van Oudenaarden

  2. Public Health Research Institute Center, New Jersey Medical School, University of Medicine and Dentistry of New Jersey, 225 Warren Street, Newark, 07103, New Jersey, USA

    Patrick van den Bogaard & Sanjay Tyagi

Authors
  1. Arjun Raj
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Patrick van den Bogaard
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Scott A Rifkin
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Alexander van Oudenaarden
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Sanjay Tyagi
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Arjun Raj or Sanjay Tyagi.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–7, Supplementary Methods (PDF 3718 kb)

Supplementary Software

Particle identification and localization software. (ZIP 3295 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Raj, A., van den Bogaard, P., Rifkin, S. et al. Imaging individual mRNA molecules using multiple singly labeled probes. Nat Methods 5, 877–879 (2008). https://doi.org/10.1038/nmeth.1253

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nmeth.1253

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing