A versatile genome-scale PCR-based pipeline for high-definition DNA FISH (original) (raw)
- Brief Communication
- Published: 23 December 2012
- Nicola Crosetto1,2,3 na1,
- Leonid Teytelman1,2,3,
- Sandy Klemm4,
- Shalev Itzkovitz1,2,3 &
- …
- Alexander van Oudenaarden1,2,3,5,6
Nature Methods volume 10, pages 122–124 (2013)Cite this article
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Abstract
We developed a cost-effective genome-scale PCR-based method for high-definition DNA FISH (HD-FISH). We visualized gene loci with diffraction-limited resolution, chromosomes as spot clusters and single genes together with transcripts by combining HD-FISH with single-molecule RNA FISH. We provide a database of over 4.3 million primer pairs targeting the human and mouse genomes that is readily usable for rapid and flexible generation of probes.
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Acknowledgements
We thank P. Junker and S. Semrau for helpful discussions. We are grateful to R.A. Weinberg (Massachusetts Institute of Technology) for providing hTERT-HME1 cells. This work was supported by the US National Institutes of Health (NIH)/National Cancer Institute Physical Sciences Oncology Center at Massachusetts Institute of Technology (U54CA143874), an NIH Pioneer award (1DP1OD003936) and a Nederlandse Organisatie voor Wetenschappelijk Onderzoek Vici award to A.v.O. M.B. and S.I. are sponsored by the Human Frontiers Science Program.
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Author notes
- Magda Bienko and Nicola Crosetto: These authors contributed equally to this work.
Authors and Affiliations
- Department of Physics, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA
Magda Bienko, Nicola Crosetto, Leonid Teytelman, Shalev Itzkovitz & Alexander van Oudenaarden - Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA
Magda Bienko, Nicola Crosetto, Leonid Teytelman, Shalev Itzkovitz & Alexander van Oudenaarden - Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA
Magda Bienko, Nicola Crosetto, Leonid Teytelman, Shalev Itzkovitz & Alexander van Oudenaarden - Department of Electrical Engineering and Computer Science, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA
Sandy Klemm - Hubrecht Institute–KNAW (Royal Netherlands Academy of Arts and Sciences), Utrecht, The Netherlands
Alexander van Oudenaarden - University Medical Center Utrecht, Utrecht, The Netherlands
Alexander van Oudenaarden
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- Magda Bienko
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Contributions
N.C. and A.v.O. conceived the methods. M.B. and N.C. performed experiments, analyzed the data and wrote the manuscript. L.T. generated the genome-wide primer databases, designed the probes and wrote the manuscript. S.K. and S.I. developed software for image processing, provided suggestions on data analysis and corrected the manuscript. A.v.O. guided experiments and data analysis, and wrote the manuscript.
Corresponding author
Correspondence toAlexander van Oudenaarden.
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The authors declare no competing financial interests.
Supplementary information
Supplementary Text and Figures
Supplementary Figures 1–5 and Supplementary Note (PDF 9821 kb)
41592_2013_BFnmeth2306_MOESM170_ESM.mov
3D animation of Chr17 in HME cells, visualized with sixteen HD-FISH probes spaced evenly every 5 Mb and labeled with two alternating fluorophores (green: AlexaFluor594; magenta: AlexaFluor647) together with a Chr17 paint probe (blue). (MOV 4960 kb)
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Bienko, M., Crosetto, N., Teytelman, L. et al. A versatile genome-scale PCR-based pipeline for high-definition DNA FISH.Nat Methods 10, 122–124 (2013). https://doi.org/10.1038/nmeth.2306
- Received: 29 August 2012
- Accepted: 14 November 2012
- Published: 23 December 2012
- Issue Date: February 2013
- DOI: https://doi.org/10.1038/nmeth.2306