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Single-cell technologies sharpen up mammalian stem cell research

Nature Cell Biology volume 16, pages 919–927 (2014)Cite this article

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Abstract

Analysis of the mechanisms underlying cell fates requires the molecular quantification of cellular features. Classical techniques use population average readouts at single time points. However, these approaches mask cellular heterogeneity and dynamics and are limited for studying rare and heterogeneous cell populations like stem cells. Techniques for single-cell analyses, ideally allowing non-invasive quantification of molecular dynamics and cellular behaviour over time, are required for studying stem cells. Here, we review the development and application of these techniques to stem cell research.

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Figure 1: Only continuous single-cell analysis can quantify molecular and cellular dynamics.
Figure 2: Single-cell technologies can reveal heterogeneities in cell populations and allow a better understanding of molecular mechanisms.
Figure 3: Examples of important conclusions from single-cell analyses.

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References

  1. Becker, A. J., McCulloch, E. A. & Till, J. E. Cytological demonstration of the clonal nature of spleen colonies derived from transplanted mouse marrow cells. Nature 197, 452–454 (1963).

    CAS  PubMed  Google Scholar 

  2. Osawa, M., Hanada, K., Hamada, H. & Nakauchi, H. Long-term lymphohematopoietic reconstitution by a single CD34-low/negative hematopoietic stem cell. Science 273, 242–245 (1996).

    CAS  PubMed  Google Scholar 

  3. Kiel, M. J. et al. SLAM family receptors distinguish hematopoietic stem and progenitor cells and reveal endothelial niches for stem cells. Cell 121, 1109–21 (2005).

    CAS  PubMed  Google Scholar 

  4. Sieburg, H. B. et al. The hematopoietic stem compartment consists of a limited number of discrete stem cell subsets. Blood 107, 2311–2316 (2006).

    CAS  PubMed  PubMed Central  Google Scholar 

  5. Dykstra, B. et al. Long-term propagation of distinct hematopoietic differentiation programs in vivo. Cell Stem Cell 1, 218–229 (2007).

    CAS  PubMed  Google Scholar 

  6. Rieger, M. A., Hoppe, P. S., Smejkal, B. M., Eitelhuber, A. C. & Schroeder, T. Hematopoietic cytokines can instruct lineage choice. Science 325, 217–218 (2009).

    CAS  PubMed  Google Scholar 

  7. Cedar, H. & Bergman, Y. Epigenetics of haematopoietic cell development. Nat. Rev. Immunol. 11, 478–488 (2011).

    CAS  PubMed  Google Scholar 

  8. Enver, T., Pera, M., Peterson, C. & Andrews, P. W. Stem cell states, fates, and the rules of attraction. Cell Stem Cell 4, 387–397 (2009).

    CAS  PubMed  Google Scholar 

  9. Moignard, V. et al. Characterization of transcriptional networks in blood stem and progenitor cells using high-throughput single-cell gene expression analysis. Nat. Cell Biol. 15, 363–372 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  10. Huang, S. Non-genetic heterogeneity of cells in development: more than just noise. Development 136, 3853–3862 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  11. Van Speybroeck, L., De Waele, D. & Van de Vijver, G. Theories in early embryology: close connections between epigenesis, preformationism, and self-organization. Ann. NY Acad. Sci. 981, 7–49 (2002).

    PubMed  Google Scholar 

  12. Schleiden, M. J. Einige Bemerkungen über den vegetabilischen Faserstoff und sein Verhältniss zum Stärkemehl. Ann. Phys. 119, 391–398 (1838).

    Google Scholar 

  13. Schwann, T. Mikroskopische Untersuchungen über die Uebereinstimmung in der Struktur und dem Wachsthum der Thiere und Pflanzen 270 (G. E. Reimer, 1839).

    Google Scholar 

  14. Virchow, R. Die Cellularpathologie in ihrer Begründung auf physiologische und pathologische Gewebelehre 582 (August Hirschwald, 1858).

    Google Scholar 

  15. Haeckel, E. Natürliche Schöpfungsgeschichte 688 (G. E. Reimer, 1868).

    Google Scholar 

  16. Pappenheim, A. Ueber Entwickelung und Ausbildung der Erythroblasten. Arch. Pathol. Anat. Physiol. Klin. Med. 145, 587–643 (1896).

    Google Scholar 

  17. Maximov, A. Der Lymphozyt als gemeinsame Stammzelle des verschiedenen Blutelemente in der embryonalen Entwicklung und im postfetalen Leben der Säugetiere. Folia Haematol. 8, 125–134 (1909).

    Google Scholar 

  18. Talbot, F. Practical Cinematography and its Applications 262 (William Heinemann, 1913).

    Google Scholar 

  19. Dunn, G. A. & Jones, G. E. Cell motility under the microscope: Vorsprung durch Technik. Nat. Rev. Mol. Cell Biol. 5, 667–672 (2004).

    CAS  PubMed  Google Scholar 

  20. Watson, J. D. & Crick, F. Molecular structure of nucleic acids: a structure for deoxyribose nucleic acid. Nature 171, 737–738 (1953).

    CAS  PubMed  Google Scholar 

  21. Pardee, A. B., Jacob, F. & Monod, J. Sur l'expression et le rôle des allèles “inductibles” et “constitutifs” dans la synthèse de la beta-galactosidase chez les zygotes d'Escherichia coli. C. R. Hebd. Seances Acad. Sci. 246, 3125–3128 (1958).

    CAS  PubMed  Google Scholar 

  22. Jacob, F. & Monod, J. Genetic regulatory mechanisms in the synthesis of proteins. J. Mol. Biol. 3, 318–356 (1961).

    CAS  PubMed  Google Scholar 

  23. Sanger, F. & Coulson, A. R. A rapid method for determining sequences in DNA by primed synthesis with DNA polymerase. J. Mol. Biol. 25, 441–448 (1975).

    Google Scholar 

  24. Saiki, R. K. et al. Enzymatic amplification of β-globin genomic sequences and restriction site analysis for diagnosis of sickle cell anemia. Science 230, 1350–1354 (1985).

    CAS  PubMed  Google Scholar 

  25. Chalfie, M., Tu, Y., Euskirchen, G., Ward, W. W. & Prasher, D. C. Green fluorescent protein as a marker for gene expression. Science 263, 802–805 (1994).

    CAS  PubMed  Google Scholar 

  26. Shaner, N. C. et al. Improved monomeric red, orange and yellow fluorescent proteins derived from Discosoma sp. red fluorescent protein. Nat. Biotechnol. 22, 1567–1572 (2004).

    CAS  PubMed  Google Scholar 

  27. Kuehn, M. R., Bradley, A., Robertson, E. J. & Evans, M. J. A potential animal model for Lesch–Nyhan syndrome through introduction of HPRT mutations into mice. Nature 326, 295–298 (1987).

    CAS  PubMed  Google Scholar 

  28. Chang, M. C. Fertilization of rabbit ova in vitro. Nature 184, 466–467 (1959).

    PubMed  Google Scholar 

  29. Uehara, T. & Yanagimachi, R. Microsurgical injection of spermatozoa into hamster eggs with subsequent transformation of sperm nuclei into male pronuclei. Biol. Reprod. 15, 467–470 (1976).

    CAS  PubMed  Google Scholar 

  30. Gardner, R. L. Mouse chimeras obtained by the injection of cells into the blastocyst. Nature 220, 596–597 (1968).

    CAS  PubMed  Google Scholar 

  31. Mintz, B. & Silvers, W. K. “Intrinsic” immunological tolerance in allophenic mice. Science 158, 1484–1486 (1967).

    CAS  PubMed  Google Scholar 

  32. Brady, G. & Iscove, N. N. Construction of cDNA libraries from single cells. Methods Enzymol. 225, 611–623 (1993).

    CAS  PubMed  Google Scholar 

  33. VanGuilder, H. D., Vrana, K. E. & Freeman, W. M. Twenty-five years of quantitative PCR for gene expression analysis. Biotechniques 44, 619–626 (2008).

    CAS  PubMed  Google Scholar 

  34. Miyamoto, T. et al. Myeloid or lymphoid promiscuity as a critical step in hematopoietic lineage commitment. Dev. Cell 3, 137–147 (2002).

    CAS  PubMed  Google Scholar 

  35. Pronk, C. J. H. et al. Elucidation of the phenotypic, functional, and molecular topography of a myeloerythroid progenitor cell hierarchy. Cell Stem Cell 1, 428–442 (2007).

    CAS  PubMed  Google Scholar 

  36. Ståhlberg, A. & Bengtsson, M. Single-cell gene expression profiling using reverse transcription quantitative real-time PCR. Methods 50, 282–288 (2010).

    PubMed  Google Scholar 

  37. Lecault, V., White, A. K., Singhal, A. & Hansen, C. L. Microfluidic single cell analysis: from promise to practice. Curr. Opin. Chem. Biol. 16, 381–390 (2012).

    CAS  PubMed  Google Scholar 

  38. Guo, G. et al. Resolution of cell fate decisions revealed by single-cell gene expression analysis from zygote to blastocyst. Dev. Cell 18, 675–685 (2010).

    CAS  PubMed  Google Scholar 

  39. Warren, L., Bryder, D., Weissman, I. L. & Quake, S. R. Transcription factor profiling in individual hematopoietic progenitors by digital RT-PCR. Proc. Natl Acad. Sci. USA 103, 17807–17812 (2006).

    CAS  PubMed  PubMed Central  Google Scholar 

  40. Hindson, B. J. et al. High-throughput droplet digital PCR system for absolute quantitation of DNA copy number. Anal. Chem. 83, 8604–8610 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Abyzov, A. et al. Somatic copy number mosaicism in human skin revealed by induced pluripotent stem cells. Nature 492, 438–442 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  42. Kurimoto, K. et al. An improved single-cell cDNA amplification method for efficient high-density oligonucleotide microarray analysis. Nucleic Acids Res. 34, e42 (2006).

    PubMed  PubMed Central  Google Scholar 

  43. Ramos, C. A. et al. Evidence for diversity in transcriptional profiles of single hematopoietic stem cells. PLoS Genet. 2, e159 (2006).

    PubMed  PubMed Central  Google Scholar 

  44. Ohnishi, Y. et al. Cell-to-cell expression variability followed by signal reinforcement progressively segregates early mouse lineages. Nat. Cell Biol. 16, 27–37 (2014).

    CAS  PubMed  Google Scholar 

  45. Wang, Z., Gerstein, M. & Snyder, M. RNA-Seq: a revolutionary tool for transcriptomics. Nat. Rev. Genet. 10, 57–63 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  46. Tang, F. et al. mRNA-Seq whole-transcriptome analysis of a single cell. Nat. Methods 6, 377–382 (2009).

    CAS  PubMed  Google Scholar 

  47. Tang, F. et al. Tracing the derivation of embryonic stem cells from the inner cell mass by single-cell RNA-Seq analysis. Cell Stem Cell 6, 468–478 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  48. Yan, L. et al. Single-cell RNA-Seq profiling of human preimplantation embryos and embryonic stem cells. Nat. Struct. Mol. Biol. 20, 1131–1139 (2013).

    CAS  PubMed  Google Scholar 

  49. Itzkovitz, S. et al. Single-molecule transcript counting of stem-cell markers in the mouse intestine. Nat. Cell Biol. 14, 106–114 (2012).

    CAS  Google Scholar 

  50. Bao, G., Rhee, W. J. & Tsourkas, A. Fluorescent probes for live-cell RNA detection. Annu. Rev. Biomed. Eng. 11, 25–47 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  51. Tyagi, S. Imaging intracellular RNA distribution and dynamics in living cells. Nat. Methods 6, 331–338 (2009).

    CAS  PubMed  Google Scholar 

  52. Bertrand, E. et al. Localization of ASH1 mRNA particles in living yeast. Mol. Cell 2, 437–445 (1998).

    CAS  PubMed  Google Scholar 

  53. Ng, K. et al. A system for imaging the regulatory noncoding Xist RNA in living mouse embryonic stem cells. Mol. Biol. Cell 22, 2634–2645 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  54. Lionnet, T. et al. A transgenic mouse for in vivo detection of endogenous labeled mRNA. Nat. Methods 8, 165–170 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  55. Larson, D. R., Zenklusen, D., Wu, B., Chao, J. A. & Singer, R. H. Real-time observation of transcription initiation and elongation on an endogenous yeast gene. Science 332, 475–478 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  56. Hocine, S., Raymond, P., Zenklusen, D., Chao, J. A. & Singer, R. H. Single-molecule analysis of gene expression using two-color RNA labeling in live yeast. Nat. Methods 10, 119–121 (2013).

    CAS  PubMed  Google Scholar 

  57. Ozawa, T., Natori, Y., Sato, M. & Umezawa, Y. Imaging dynamics of endogenous mitochondrial RNA in single living cells. Nat. Methods 4, 413–419 (2007).

    CAS  PubMed  Google Scholar 

  58. Bendall, S. C. et al. Single-cell mass cytometry of differential immune and drug responses across a human hematopoietic continuum. Science 332, 687–696 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  59. Newell, E. W., Sigal, N., Bendall, S. C., Nolan, G. P. & Davis, M. M. Cytometry by time-of-flight shows combinatorial cytokine expression and virus-specific cell niches within a continuum of CD8+ T cell phenotypes. Immunity 36, 142–152 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  60. Qiu, P. et al. Extracting a cellular hierarchy from high-dimensional cytometry data with SPADE. Nat. Biotechnol. 29, 886–891 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  61. Ornatsky, O. et al. Highly multiparametric analysis by mass cytometry. J. Immunol. Methods 361, 1–20 (2010).

    CAS  PubMed  Google Scholar 

  62. Han, L. et al. Single-cell mass cytometry reveals phenotypic and functional heterogeneity in acute myeloid leukemia at diagnosis and in remission. Blood 122, 1311 (2013).

    Google Scholar 

  63. Beckervordersandforth, R. et al. In vivo fate mapping and expression analysis reveals molecular hallmarks of prospectively isolated adult neural stem cells. Cell Stem Cell 7, 744–758 (2010).

    CAS  PubMed  Google Scholar 

  64. Barker, N. et al. Identification of stem cells in small intestine and colon by marker gene Lgr5. Nature 449, 1003–1007 (2007).

    CAS  PubMed  Google Scholar 

  65. Park, D. et al. Endogenous bone marrow MSCs are dynamic, fate-restricted participants in bone maintenance and regeneration. Cell Stem Cell 10, 259–272 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  66. Ortega, F. et al. Oligodendrogliogenic and neurogenic adult subependymal zone neural stem cells constitute distinct lineages and exhibit differential responsiveness to Wnt signalling. Nat. Cell Biol. 15, 602–613 (2013).

    CAS  PubMed  Google Scholar 

  67. Zuba-Surma, E. K. & Ratajczak, M. Z. Analytical capabilities of the ImageStream cytometer. Methods Cell Biol. 102, 207–230 (2011).

    PubMed  Google Scholar 

  68. Lapid, K. et al. GSK3β regulates physiological migration of stem/progenitor cells via cytoskeletal rearrangement. J. Clin. Invest. 123, 1705–1717 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  69. Garini, Y., Young, I. T. & Mcnamara, G. Spectral imaging: principles and applications. Cytometry Part A 69, 735–747 (2006).

    Google Scholar 

  70. Zimmermann, T. Spectral imaging and linear unmixing in light microscopy. Adv. Biochem. Eng. Biotechnol. 95, 245–265 (2005).

    PubMed  Google Scholar 

  71. Rothbauer, U. et al. Targeting and tracing antigens in live cells with fluorescent nanobodies. Nat. Methods 3, 887–889 (2006).

    CAS  PubMed  Google Scholar 

  72. Chambers, I. et al. Nanog safeguards pluripotency and mediates germline development. Nature 450, 1230–1234 (2007).

    CAS  PubMed  Google Scholar 

  73. Filipczyk, A. et al. Biallelic expression of nanog protein in mouse embryonic stem cells. Cell Stem Cell 13, 12–13 (2013).

    CAS  PubMed  Google Scholar 

  74. Wang, L. et al. Identification of a clonally expanding haematopoietic compartment in bone marrow. EMBO J. 32, 219–230 (2013).

    PubMed  Google Scholar 

  75. Jordan, C. T. & Lemischka, I. R. Clonal and systemic analysis of long-term hematopoiesis in the mouse. Genes Dev. 4, 220–232 (1990).

    CAS  PubMed  Google Scholar 

  76. Mazurier, F., Gan, O. I., McKenzie, J. L., Doedens, M. & Dick, J. E. Lentivector-mediated clonal tracking reveals intrinsic heterogeneity in the human hematopoietic stem cell compartment and culture-induced stem cell impairment. Blood 103, 545–552 (2004).

    CAS  PubMed  Google Scholar 

  77. Ema, H. et al. Adult mouse hematopoietic stem cells: purification and single-cell assays. Nat. Protoc. 1, 2979–2987 (2006).

    CAS  PubMed  Google Scholar 

  78. Tzouanacou, E., Wegener, A., Wymeersch, F. J., Wilson, V. & Nicolas, J.-F. Redefining the progression of lineage segregations during mammalian embryogenesis by clonal analysis. Dev. Cell 17, 365–376 (2009).

    CAS  PubMed  Google Scholar 

  79. Beckervordersandforth, R. et al. In vivo targeting of adult neural stem cells in the dentate gyrus by a split-Cre approach. Stem Cell Reports 2, 153–162 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  80. Buczacki, S. J. a. et al. Intestinal label-retaining cells are secretory precursors expressing Lgr5. Nature 495, 65–69 (2013).

    CAS  PubMed  Google Scholar 

  81. Livet, J. et al. Transgenic strategies for combinatorial expression of fluorescent proteins in the nervous system. Nature 450, 56–62 (2007).

    CAS  PubMed  Google Scholar 

  82. Snippert, H. J. et al. Intestinal crypt homeostasis results from neutral competition between symmetrically dividing Lgr5 stem cells. Cell 143, 134–144 (2010).

    CAS  PubMed  Google Scholar 

  83. Schepers, A. G. et al. Lineage tracing reveals Lgr5+ stem cell activity in mouse intestinal adenomas. Science 337, 730–735 (2012).

    CAS  PubMed  Google Scholar 

  84. Cai, D., Cohen, K. B., Luo, T., Lichtman, J. W. & Sanes, J. R. Improved tools for the Brainbow toolbox. Nat. Methods 10, 540–547 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  85. Tasic, B. et al. Extensions of MADM (mosaic analysis with double markers) in mice. PLoS ONE 7, e33332 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  86. Espinosa, J. S. & Luo, L. Timing Neurogenesis and differentiation: Insights from quantitative clonal analyses of cerebellar granule cells. J. Neurosci. 28, 2301–2312 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  87. Espinosa, J. S., Wheeler, D. G., Tsien, R. W. & Luo, L. Uncoupling dendrite growth and patterning: single-cell knockout analysis of NMDA receptor 2B. Neuron 62, 205–217 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  88. Hippenmeyer, S. et al. Genetic mosaic dissection of Lis1 and Ndel1 in neuronal migration. Neuron 68, 695–709 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  89. Liu, C. et al. Mosaic analysis with double markers reveals tumor cell of origin in glioma. Cell 146, 209–221 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  90. Lu, R., Neff, N. F., Quake, S. R. & Weissman, I. L. Tracking single hematopoietic stem cells in vivo using high-throughput sequencing in conjunction with viral genetic barcoding. Nat. Biotechnol. 29, 928–933 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  91. Gerrits, A. et al. Cellular barcoding tool for clonal analysis in the hematopoietic system. Blood 115, 2610–2618 (2010).

    CAS  PubMed  Google Scholar 

  92. Verovskaya, E. et al. Heterogeneity of young and aged murine hematopoietic stem cells revealed by quantitative clonal analysis using cellular barcoding. Blood 122, 523–532 (2013).

    CAS  PubMed  Google Scholar 

  93. Schepers, K. et al. Dissecting T cell lineage relationships by cellular barcoding. J. Exp. Med. 205, 2309–2318 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  94. Naik, S. H. et al. Diverse and heritable lineage imprinting of early haematopoietic progenitors. Nature 496, 229–232 (2013).

    CAS  PubMed  Google Scholar 

  95. Malide, D., Métais, J.-Y. & Dunbar, C. E. Dynamic clonal analysis of murine hematopoietic stem and progenitor cells marked by 5 fluorescent proteins using confocal and multiphoton microscopy. Blood 120, e105–116 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  96. Coutu, D. L. & Schroeder, T. Probing cellular processes by long-term live imaging—historic problems and current solutions. J. Cell Sci. 126, 3805–3815 (2013).

    CAS  PubMed  Google Scholar 

  97. Kuang, S., Kuroda, K., Le Grand, F. & Rudnicki, M. A. Asymmetric self-renewal and commitment of satellite stem cells in muscle. Cell 129, 999–1010 (2007).

    CAS  PubMed  PubMed Central  Google Scholar 

  98. Burtscher, I. & Lickert, H. Foxa2 regulates polarity and epithelialization in the endoderm germ layer of the mouse embryo. Development 136, 1029–1038 (2009).

    CAS  PubMed  Google Scholar 

  99. Eilken, H. M., Nishikawa, S.-I. & Schroeder, T. Continuous single-cell imaging of blood generation from haemogenic endothelium. Nature 457, 896–900 (2009).

    CAS  PubMed  Google Scholar 

  100. Bertrand, J. Y. et al. Haematopoietic stem cells derive directly from aortic endothelium during development. Nature 464, 108–111 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  101. Kissa, K. & Herbomel, P. Blood stem cells emerge from aortic endothelium by a novel type of cell transition. Nature 464, 112–115 (2010).

    CAS  PubMed  Google Scholar 

  102. Asami, M. et al. The role of Pax6 in regulating the orientation and mode of cell division of progenitors in the mouse cerebral cortex. Development 138, 5067–5078 (2011).

    CAS  PubMed  Google Scholar 

  103. Costa, M. R., Wen, G., Lepier, A., Schroeder, T. & Götz, M. Par-complex proteins promote proliferative progenitor divisions in the developing mouse cerebral cortex. Development 135, 11–22 (2008).

    CAS  PubMed  Google Scholar 

  104. Costa, M. R., Bucholz, O., Schroeder, T. & Götz, M. Late origin of glia-restricted progenitors in the developing mouse cerebral cortex. Cereb. Cortex 19 (suppl. 1), i135–i143 (2009).

    PubMed  Google Scholar 

  105. Costa, M. R. et al. Continuous live imaging of adult neural stem cell division and lineage progression in vitro. Development 138, 1057–1068 (2011).

    CAS  PubMed  Google Scholar 

  106. Rompolas, P. et al. Live imaging of stem cell and progeny behaviour in physiological hair-follicle regeneration. Nature 487, 496–499 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  107. Ritsma, L. et al. Intestinal crypt homeostasis revealed at single-stem-cell level by in vivo live imaging. Nature 507, 362–365 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  108. Wu, M. & Singh, A. K. Single-cell protein analysis. Curr. Opin. Biotechnol. 23, 83–88 (2012).

    CAS  PubMed  Google Scholar 

  109. Ståhlberg, A., Rusnakova, V., Forootan, A., Anderova, M. & Kubista, M. RT-qPCR work-flow for single-cell data analysis. Methods 59, 80–88 (2013).

    PubMed  Google Scholar 

  110. Tang, F., Lao, K. & Surani, M. A. Development and applications of single-cell transcriptome analysis. Nat. Methods 8, S6–S11 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  111. Shapiro, E., Biezuner, T. & Linnarsson, S. Single-cell sequencing-based technologies will revolutionize whole-organism science. Nat. Rev. Genet. 14, 618–630 (2013).

    CAS  PubMed  Google Scholar 

  112. Schadt, E. E., Turner, S. & Kasarskis, A. A window into third-generation sequencing. Hum. Mol. Genet. 19, R227–R240 (2010).

    CAS  PubMed  Google Scholar 

  113. Femino, A. M., Fay, F. S., Fogarty, K. & Singer, R. H. Visualization of single RNA transcripts in situ. Science 280, 585–590 (1998).

    CAS  PubMed  Google Scholar 

  114. Itzkovitz, S. & van Oudenaarden, A. Validating transcripts with probes and imaging technology. Nat. Methods 8, S12–S19 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  115. Levsky, J. M., Shenoy, S. M., Pezo, R. C. & Singer, R. H. Single-cell gene expression profiling. Science 297, 836–840 (2002).

    CAS  PubMed  Google Scholar 

  116. Raj, A., van den Bogaard, P., Rifkin, S. A., van Oudenaarden, A. & Tyagi, S. Imaging individual mRNA molecules using multiple singly labeled probes. Nat. Methods 5, 877–879 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  117. Tyagi, S. & Kramer, F. R. Molecular beacons: probes that fluoresce upon hybridization. Nat. Biotechnol. 14, 303–308 (1996).

    CAS  PubMed  Google Scholar 

  118. Santangelo, P. J., Nix, B., Tsourkas, A. & Bao, G. Dual FRET molecular beacons for mRNA detection in living cells. Nucleic Acids Res. 32, e57 (2004).

    PubMed  PubMed Central  Google Scholar 

  119. Bratu, D. P., Cha, B.-J., Mhlanga, M. M., Kramer, F. R. & Tyagi, S. Visualizing the distribution and transport of mRNAs in living cells. Proc. Natl Acad. Sci. USA 100, 13308–13313 (2003).

    CAS  PubMed  PubMed Central  Google Scholar 

  120. Paige, J. S., Wu, K. Y. & Jaffrey, S. R. RNA mimics of green fluorescent protein. Science 333, 642–646 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  121. Seferos, D. S., Giljohann, D. A., Hill, H. D., Prigodich, A. E. & Mirkin, C. A. Nano-flares: probes for transfection and mRNA detection in living cells. J. Am. Chem. Soc. 129, 15477–15479 (2007).

    CAS  PubMed  PubMed Central  Google Scholar 

  122. Hu, C.-D. & Kerppola, T. K. Simultaneous visualization of multiple protein interactions in living cells using multicolor fluorescence complementation analysis. Nat. Biotechnol. 21, 539–545 (2003).

    CAS  PubMed  PubMed Central  Google Scholar 

  123. Kerppola, T. K. Complementary methods for studies of protein interactions in living cells. Nat. Methods 3, 969–971 (2006).

    CAS  PubMed  PubMed Central  Google Scholar 

  124. Herce, H. D., Deng, W., Helma, J., Leonhardt, H. & Cardoso, M. C. Visualization and targeted disruption of protein interactions in living cells. Nat. Commun. 4, 2660 (2013).

    PubMed  Google Scholar 

  125. Gomez, D., Shankman, L. S., Nguyen, A. T. & Owens, G. K. Detection of histone modifications at specific gene loci in single cells in histological sections. Nat. Methods 10, 171–177 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  126. Zhang, B. et al. Microenvironmental protection of CML stem and progenitor cells from tyrosine kinase inhibitors through N-cadherin and Wnt-β-catenin signaling. Blood 121, 1824–1838 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  127. Endele, M. & Schroeder, T. Molecular live cell bioimaging in stem cell research. Ann. NY Acad. Sci. 1266, 18–27 (2012).

    CAS  PubMed  Google Scholar 

  128. Newman, R., Fosbrink, M. & Zhang, J. Genetically encodable fluorescent biosensors for tracking signaling dynamics in living cells. Chem. Rev. 111, 3614–3666 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  129. Jaisser, F. Inducible gene expression and gene modification in transgenic mice. J. Am. Soc. Nephrol. 11 (suppl. 1), S95–S100 (2000).

    CAS  PubMed  Google Scholar 

  130. Abe, T. et al. Establishment of conditional reporter mouse lines at ROSA26 locus for live cell imaging. Genesis 49, 579–590 (2011).

    CAS  PubMed  Google Scholar 

  131. Kühn, R., Schwenk, F., Aguet, M. & Rajewsky, K. Inducible gene targeting in mice. Science 269, 1427–1429 (1995).

    PubMed  Google Scholar 

  132. Banaszynski, L., Chen, L., Maynard-Smith, L. A., Ooi, G. L. & Wandless, T. J. A rapid, reversible, and tunable method to regulate protein function in living cells using synthetic small molecules. Cell 126, 995–1004 (2006).

    CAS  PubMed  PubMed Central  Google Scholar 

  133. Banaszynski, L. A., Sellmyer, M. A., Contag, C. H., Wandless, T. J. & Thorne, S. H. Chemical control of protein stability and function in living mice. Nat. Med. 14, 1123–1127 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  134. Bacchus, W. & Fussenegger, M. The use of light for engineered control and reprogramming of cellular functions. Curr. Opin. Biotechnol. 23, 695–702 (2012).

    CAS  PubMed  Google Scholar 

  135. Ye, H., Daoud-El Baba, M., Peng, R.-W. & Fussenegger, M. A synthetic optogenetic transcription device enhances blood-glucose homeostasis in mice. Science 332, 1565–1568 (2011).

    CAS  PubMed  Google Scholar 

  136. Moreau-Gachelin, F., Tavitian, A. & Tambourin, P. Spi-1 is a putative oncogene in virally induced murine erythroleukaemias. Nature 331, 277–280 (1988).

    CAS  PubMed  Google Scholar 

  137. Iwama, A. et al. Use of RDA analysis of knockout mice to identify myeloid genes regulated in vivo by PU.1 and C/EBPα. Nucleic Acids Res. 26, 3034–3043 (1998).

    CAS  PubMed  PubMed Central  Google Scholar 

  138. Akashi, K., Traver, D., Miyamoto, T. & Weissman, I. L. A clonogenic common myeloid progenitor that gives rise to all myeloid lineages. Nature 404, 193–197 (2000).

    CAS  PubMed  Google Scholar 

  139. Arinobu, Y. et al. Reciprocal activation of GATA-1 and PU.1 marks initial specification of hematopoietic stem cells into myeloerythroid and myelolymphoid lineages. Cell Stem Cell 1, 416–427 (2007).

    CAS  PubMed  Google Scholar 

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  1. Department of Biosystems Science and Engineering, ETH Zurich, Basel, 4058, Switzerland

    Philipp S. Hoppe, Daniel L. Coutu & Timm Schroeder

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  1. Philipp S. Hoppe
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  2. Daniel L. Coutu
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Correspondence to Timm Schroeder.

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Hoppe, P., Coutu, D. & Schroeder, T. Single-cell technologies sharpen up mammalian stem cell research. Nat Cell Biol 16, 919–927 (2014). https://doi.org/10.1038/ncb3042

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