Allele-specific detection of single mrna molecules in situ

Allele-specific detection of single mrna molecules in situ


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ABSTRACT We describe a method for fluorescence _in situ_ identification of individual mRNA molecules, allowing quantitative and accurate measurement, in single cells, of allele-specific


transcripts that differ by only a few nucleotides. By using a combination of allele-specific and non–allele-specific probe libraries, we achieve >95% detection accuracy. We investigate


the allele-specific stochastic expression of _Nanog_, which encodes a pluripotency factor, in murine embryonic stem cells. Access through your institution Buy or subscribe This is a preview


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SINGLE-CELL LINEAGE TRACING IN MICE AND HUMANS BASED ON DNA METHYLATION EPIMUTATIONS Article 16 January 2025 SINGLE-MOLECULE IMAGING OF TRANSCRIPTION DYNAMICS IN SOMATIC STEM CELLS Article


24 June 2020 IFLPMOSAICS ENABLE THE MULTISPECTRAL BARCODING AND HIGH-THROUGHPUT COMPARATIVE ANALYSIS OF MUTANT AND WILD-TYPE CELLS Article Open access 13 December 2024 REFERENCES * Eldar, A.


& Elowitz, M.B. _Nature_ 467, 167–173 (2010). Article  CAS  Google Scholar  * Balázsi, G., van Oudenaarden, A. & Collins, J.J. _Cell_ 144, 910–925 (2011). Article  Google Scholar  *


Femino, A.M., Fay, F.S., Fogarty, K. & Singer, R.H. _Science_ 280, 585–590 (1998). Article  CAS  Google Scholar  * Raj, A., van den Bogaard, P., Rifkin, S.A., van Oudenaarden, A. &


Tyagi, S. _Nat. Methods_ 5, 877–879 (2008). Article  CAS  Google Scholar  * White, A.K. et al. _Proc. Natl. Acad. Sci. USA_ 108, 13999–14004 (2011). Article  CAS  Google Scholar  * Larsson,


C., Grundberg, I., Söderberg, O. & Nilsson, M. _Nat. Methods_ 7, 395–397 (2010). Article  CAS  Google Scholar  * Keane, T.M. et al. _Nature_ 477, 289–294 (2011). Article  CAS  Google


Scholar  * Panning, B., Dausman, J. & Jaenisch, R. _Cell_ 90, 907–916 (1997). Article  CAS  Google Scholar  * Yilmaz, L.S., Parkernar, S. & Noguera, D.R. _Appl. Environ. Microbiol._


77, 1118–1122 (2011). Article  CAS  Google Scholar  * Ying, Q.L. et al. _Nature_ 453, 519–523 (2008). Article  CAS  Google Scholar  * Miyanari, Y. & Torres-Padilla, M.E. _Nature_ 483,


470–473 (2012). Article  CAS  Google Scholar  * Levesque, M.J. & Raj, A. _Nat. Methods_ 10, 246–248 (2013). Article  CAS  Google Scholar  * Raj, A., Peskin, C.S., Tranchina, D., Vargas,


D.Y. & Tyagi, S. _PLoS Biol._ 4, e309 (2006). Article  Google Scholar  * Zhong, J.F. et al. _Lab Chip_ 8, 68–74 (2008). Article  CAS  Google Scholar  Download references ACKNOWLEDGEMENTS


We thank S. Semrau for programming assistance and B. Panning (University of California, San Francisco) for providing the 2-1 embryonic stem cells and helpful discussions and for inspiring


the initial idea that led to the development of this method. We thank J.P. Junker, S. Klemm, Y. Lin, D. Mooijman, A. Pawlosky and Y. Zheng for technical assistance, general discussions


and/or comments on the manuscript. This work was supported by a grant (U54CA143874) from the US National Institutes of Health (NIH) National Cancer Institute Physical Sciences Oncology


Center at Massachusetts Institute of Technology, an NIH Pioneer Award (8 DP1 CA174420-05) and a Nederlandse Organisatie voor Wetenschappelijk Onderzoek (NWO) Vici award to A.v.O. AUTHOR


INFORMATION AUTHORS AND AFFILIATIONS * Harvard University Graduate Biophysics Program, Harvard Medical School, Boston, Massachusetts, USA Clinton H Hansen * Department of Physics,


Massachusetts Institute of Technology, Cambridge, Massachusetts, USA Alexander van Oudenaarden * Department of Biology, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA


Alexander van Oudenaarden * Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA Alexander van Oudenaarden * Hubrecht


Institute—Royal Netherlands Academy of Arts and Sciences (KNAW), University Medical Center Utrecht, Utrecht, The Netherlands Alexander van Oudenaarden Authors * Clinton H Hansen View author


publications You can also search for this author inPubMed Google Scholar * Alexander van Oudenaarden View author publications You can also search for this author inPubMed Google Scholar


CONTRIBUTIONS C.H.H. and A.v.O. conceived the method. C.H.H. performed experiments, analyzed the data and wrote the manuscript. A.v.O. guided experiments and data analysis and wrote the


manuscript. CORRESPONDING AUTHOR Correspondence to Alexander van Oudenaarden. ETHICS DECLARATIONS COMPETING INTERESTS The authors declare no competing financial interests. SUPPLEMENTARY


INFORMATION SUPPLEMENTARY TEXT AND FIGURES Supplementary Figures 1–18, Supplementary Table 2 and Supplementary Note (PDF 1768 kb) SUPPLEMENTARY TABLE 1 Probe Sequences (TXT 27 kb) RIGHTS AND


PERMISSIONS Reprints and permissions ABOUT THIS ARTICLE CITE THIS ARTICLE Hansen, C., van Oudenaarden, A. Allele-specific detection of single mRNA molecules _in situ_. _Nat Methods_ 10,


869–871 (2013). https://doi.org/10.1038/nmeth.2601 Download citation * Received: 19 April 2013 * Accepted: 19 July 2013 * Published: 11 August 2013 * Issue Date: September 2013 * DOI:


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