Rgb marking facilitates multicolor clonal cell tracking

ABSTRACT We simultaneously transduced cells with three lentiviral gene ontology (LeGO) vectors encoding red, green or blue fluorescent proteins. Individual cells were thereby marked by

different combinations of inserted vectors, resulting in the generation of numerous mixed colors, a principle we named red-green-blue (RGB) marking. We show that lentiviral vector–mediated

RGB marking remained stable after cell division, thus facilitating the analysis of clonal cell fates _in vitro_ and _in vivo_. Particularly, we provide evidence that RGB marking allows

assessment of clonality after regeneration of injured livers by transplanted primary hepatocytes. We also used RGB vectors to mark hematopoietic stem/progenitor cells that generated colored

spleen colonies. Finally, based on limiting-dilution and serial transplantation assays with tumor cells, we found that clonal tumor cells retained their specific color-code over extensive

periods of time. We conclude that RGB marking represents a useful tool for cell clonality studies in tissue regeneration and pathology. Access through your institution Buy or subscribe This

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OF CLONAL TRACKING DATA USING BARCODETRACKR Article 26 April 2021 IFLPMOSAICS ENABLE THE MULTISPECTRAL BARCODING AND HIGH-THROUGHPUT COMPARATIVE ANALYSIS OF MUTANT AND WILD-TYPE CELLS

Article Open access 13 December 2024 ISOLATING LIVE CELL CLONES FROM BARCODED POPULATIONS USING CRISPRA-INDUCIBLE REPORTERS Article 27 July 2020 REFERENCES * Shaner, N.C., Steinbach, P.

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_Methods Mol. Biol._ 132, 365–386 (2000). CAS  Google Scholar  Download references ACKNOWLEDGEMENTS We wish to thank to V. Matzat and U. Bergholz for expert technical assistance, R. Reusch

for excellent mouse care and J. Petersen for continuous support with the uPA model and critical reading of the manuscript. Confocal imaging was performed with the kind help from O. Bruns in

collaboration with the Nikon Application Center Northern Germany (Nikon). We are indebted to many colleagues for their kind support with various cells and constructs: H. Wege (University

Medical Center Hamburg-Eppendorf) for FH-hTERT, D. Hösch (University Marburg) for BON cells, W. Beyer (Heinrich-Pette-Institute) for vesicular stomatitis virus G protein cDNA R.Y. Tsien

(Howard Hughes Medical Institute) for mCherry cDNA, A. Miyawaki (RIKEN) and T. Schroeder (Institute for Stem Cell Research) for Venus cDNA and D.W. Piston (Vanderbilt-Ingram Cancer Center)

for Cerulean cDNA. This work was supported by the Deutsche Forschungsgemeinschaft (SFB841 to B.F., D.B. and M.D. and FE568/11-1 to B.F.). M. Thomaschewski was supported by the integrated

graduate school “Inflammation and Regeneration” of the University Medical Center Hamburg-Eppendorf. AUTHOR INFORMATION AUTHORS AND AFFILIATIONS * Research Department Cell and Gene Therapy,

Clinic for Stem Cell Transplantation, University Cancer Center Hamburg, University Medical Center Hamburg-Eppendorf, Hamburg, Germany Kristoffer Weber, Michael Thomaschewski, Michael

Warlich, Kerstin Cornils & Boris Fehse * Internal Medicine I, University Medical Center Hamburg-Eppendorf, Hamburg, Germany Michael Warlich, Tassilo Volz, Marc Lütgehetmann, Maura Dandri

 & Daniel Benten * Heinrich-Pette-Institute, Leibniz-Institute for Experimental Virology, Hamburg, Germany Birte Niebuhr, Maike Täger & Carol Stocking * Department of Hepatobiliary

and Transplant Surgery, University Medical Center Hamburg-Eppendorf, Hamburg, Germany Jörg-Matthias Pollok Authors * Kristoffer Weber View author publications You can also search for this

author inPubMed Google Scholar * Michael Thomaschewski View author publications You can also search for this author inPubMed Google Scholar * Michael Warlich View author publications You can

also search for this author inPubMed Google Scholar * Tassilo Volz View author publications You can also search for this author inPubMed Google Scholar * Kerstin Cornils View author

publications You can also search for this author inPubMed Google Scholar * Birte Niebuhr View author publications You can also search for this author inPubMed Google Scholar * Maike Täger

View author publications You can also search for this author inPubMed Google Scholar * Marc Lütgehetmann View author publications You can also search for this author inPubMed Google Scholar

* Jörg-Matthias Pollok View author publications You can also search for this author inPubMed Google Scholar * Carol Stocking View author publications You can also search for this author

inPubMed Google Scholar * Maura Dandri View author publications You can also search for this author inPubMed Google Scholar * Daniel Benten View author publications You can also search for

this author inPubMed Google Scholar * Boris Fehse View author publications You can also search for this author inPubMed Google Scholar CONTRIBUTIONS K.W. designed the study, produced LeGO

vectors and performed and analyzed gene transfer experiments in target cells _in vitro_ and _in vivo_. M. Thomaschewski and M.W. isolated, transduced and transplanted primary hepatocytes and

analyzed mice. M. Thomaschewski also performed mouse studies with BON tumor cells. T.V. and M.L. performed experiments in uPA mice. K.C. identified vector insertions by LM-PCR and performed

specific PCRs. B.N., M. Täger and C.S. designed and performed experiments with mouse HSCs. J.-M.P. and M.L. provided and prepared primary human hepatocytes. M.D. provided the uPA-SCID model

and supervised _in vivo_ experiments in that model. D.B. designed and performed mouse studies. B.F. designed the study, analyzed and evaluated results and wrote the manuscript. All authors

read and approved the final version of the manuscript. CORRESPONDING AUTHOR Correspondence to Boris Fehse. ETHICS DECLARATIONS COMPETING INTERESTS The authors declare no competing financial

interests. SUPPLEMENTARY INFORMATION SUPPLEMENTARY TEXT AND FIGURES Supplementary Figures 1–4 and Supplementary Tables 1 and 2 (PDF 662 kb) RIGHTS AND PERMISSIONS Reprints and permissions

ABOUT THIS ARTICLE CITE THIS ARTICLE Weber, K., Thomaschewski, M., Warlich, M. _et al._ RGB marking facilitates multicolor clonal cell tracking. _Nat Med_ 17, 504–509 (2011).

https://doi.org/10.1038/nm.2338 Download citation * Received: 10 May 2010 * Accepted: 01 December 2010 * Published: 27 March 2011 * Issue Date: April 2011 * DOI:

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