Brief Communication | Published:

In situ sequencing for RNA analysis in preserved tissue and cells

Nature Methods volume 10, pages 857860 (2013) | Download Citation

Abstract

Tissue gene expression profiling is performed on homogenates or on populations of isolated single cells to resolve molecular states of different cell types. In both approaches, histological context is lost. We have developed an in situ sequencing method for parallel targeted analysis of short RNA fragments in morphologically preserved cells and tissue. We demonstrate in situ sequencing of point mutations and multiplexed gene expression profiling in human breast cancer tissue sections.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

References

  1. 1.

    & Trends Cell Biol. 13, 4–6 (2003).

  2. 2.

    , & Nat. Rev. Genet. 10, 57–63 (2009).

  3. 3.

    et al. Science 278, 1481–1483 (1997).

  4. 4.

    et al. Nat. Biotechnol. 29, 1120–1127 (2011).

  5. 5.

    et al. Nature 472, 90–94 (2011).

  6. 6.

    et al. Nat. Methods 6, 377–382 (2009).

  7. 7.

    et al. Cell 148, 873–885 (2012).

  8. 8.

    et al. Cell 148, 886–895 (2012).

  9. 9.

    et al. Science 265, 2085–2088 (1994).

  10. 10.

    , , & Nucleic Acids Res. 26, 5073–5078 (1998).

  11. 11.

    , , & Nat. Methods 7, 395–397 (2010).

  12. 12.

    et al. Science 309, 1728–1732 (2005).

  13. 13.

    et al. Science 327, 78–81 (2010).

  14. 14.

    et al. Bioinformatics 27, 1179–1180 (2011).

  15. 15.

    , & IEEE Trans. Image Process. 7, 27–41 (1998).

  16. 16.

    & J. Clin. Oncol. 26, 721–728 (2008).

  17. 17.

    et al. Nature 456, 470–476 (2008).

Download references

Acknowledgements

We thank J. Lee, G.M. Church and F. Pontén for valuable discussion about this work. We thank M. Dahlberg for helping extracting RNA-seq data from publications. The research reported in this paper was funded by the Swedish Research Council, VINNOVA project “Companion diagnostic initiative,” the European Community's 7th Framework Program (FP7/2007-2013) under grant agreement nos. 259796 (DiaTools) and 201418 (READNA), the Science for Life Laboratory, Stockholm and Uppsala, and the Innovative Medicines Initiative Joint Undertaking under grant agreement no. 115234 (OncoTrack).

Author information

Author notes

    • Rongqin Ke
    •  & Marco Mignardi

    These authors contributed equally to this work.

Affiliations

  1. Science for Life Laboratory, Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden.

    • Rongqin Ke
    • , Marco Mignardi
    • , Jessica Svedlund
    •  & Mats Nilsson
  2. Department of Immunology, Genetics, and Pathology, the Rudbeck Laboratory, Uppsala University, Uppsala, Sweden.

    • Rongqin Ke
    • , Marco Mignardi
    • , Johan Botling
    •  & Mats Nilsson
  3. Science for Life Laboratory, Centre for Image Analysis, Uppsala University, Uppsala, Sweden.

    • Alexandra Pacureanu
    •  & Carolina Wählby
  4. Imaging Platform, Broad Institute of Harvard and Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.

    • Carolina Wählby

Authors

  1. Search for Rongqin Ke in:

  2. Search for Marco Mignardi in:

  3. Search for Alexandra Pacureanu in:

  4. Search for Jessica Svedlund in:

  5. Search for Johan Botling in:

  6. Search for Carolina Wählby in:

  7. Search for Mats Nilsson in:

Contributions

R.K. and M.M. designed and performed the experiments. J.B. provided tissue sections and pathology examination of the tissue. C.W. designed the image analysis pipelines and performed the image analysis together with A.P. J.S. performed the correlation between in situ sequencing and RNA-seq data. R.K., M.M., C.W. and M.N. wrote the manuscript. All authors commented on and revised the manuscript. M.N. conceived the idea and supervised the project.

Competing interests

M.N. owns shares in the company Olink AB, Uppsala, Sweden, which holds patents whose value may be affected by publication of these results.

Corresponding authors

Correspondence to Carolina Wählby or Mats Nilsson.

Supplementary information

PDF files

  1. 1.

    Supplementary Text and Figures

    Supplementary Figures 1–15, Supplementary Tables 1–7 and Supplementary Notes 1 and 2

About this article

Publication history

Received

Accepted

Published

DOI

https://doi.org/10.1038/nmeth.2563

Further reading