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pp 1-13 | Cite as

Tracking Germline Stem Cell Dynamics In Vivo in C. elegans Using Photoconversion

  • Simona Rosu
  • Orna Cohen-FixEmail author
Protocol
Part of the Methods in Molecular Biology book series

Abstract

The maintenance of many adult tissues depends on stem cell systems, which must balance proliferation and differentiation. To understand the properties of adult stem cell systems, one powerful tool is visualization of the cell dynamics in vivo. Here we describe a protocol to track cells in the germline progenitor zone (which includes germline stem cells) in live C. elegans adult worms. Tracking is achieved by using a genetically encoded photoconvertible fluorescent protein, where photoconversion is used to mark cells of interest and their descendants. Individual worms are immobilized, the cells of interest are selected for photoconversion, and the worms are then recovered to plates and imaged at later timepoints. This allows longitudinal studies of individual worms, providing valuable information regarding germline stem cell dynamics.

Keywords

Germline Stem cells Photoconversion Proliferation C. elegans Lineage tracing Live imaging 

Notes

Acknowledgments

The authors were supported by the Intramural Research Program of the NIH, The National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK).

References

  1. 1.
    Rosu S, Cohen-Fix O (2017) Live-imaging analysis of germ cell proliferation in the C. elegans adult supports a stochastic model for stem cell proliferation. Dev Biol 423(2):93–100Google Scholar
  2. 2.
    Lukyanov KA, Chudakov DM, Lukyanov S, Verkhusha VV (2005) Photoactivatable fluorescent proteins. Nat Rev Mol Cell Biol 6:885–890Google Scholar
  3. 3.
    Gurskaya NG, Verkhusha VV, Shcheglov AS, Staroverov DB, Chepurnykh TV, Fradkov AF, Lukyanov S, Lukyanov KA (2006) Engineering of a monomeric green-to-red photoactivatable fluorescent protein induced by blue light. Nat Biotechnol 24:461–465Google Scholar
  4. 4.
    Dempsey WP, Georgieva L, Helbling PM, Sonay AY, Truong TV, Haffner M, Pantazis P (2015) In vivo single-cell labeling by confined primed conversion. Nat Methods 12:645–648Google Scholar
  5. 5.
    Ivanchenko S, Glaschick S, Rocker C, Oswald F, Wiedenmann J, Ulrich N (2007) Two-photon excitation and photoconversion of EosFP in dual-color 4Pi confocal microscopy. Biophys J 92(12):4451–4457Google Scholar
  6. 6.
    McCarter J, Bartlett B, Dang T, Schedl T (1999) On the control of oocyte meiotic maturation and ovulation in Caenorhabditis elegans. Dev Biol 205:111–128Google Scholar
  7. 7.
    Kim E, Sun L, Gabel CV, Fang-Yen C (2013) Long-term imaging of Caenorhabditis elegans using nanoparticle-mediated immobilization. PLoS One 8:e53419Google Scholar
  8. 8.
    Chudakov DM, Lukyanov S, Lukyanov KA (2007) Tracking intracellular protein movements using photoswitchable fluorescent proteins PS-CFP2 and Dendra2. Nat Protoc 2:2024–2032Google Scholar

Copyright information

© Springer Science+Business Media New York 2019

Authors and Affiliations

  1. 1.The Graduate SchoolSyracuse UniversitySyracuseUSA
  2. 2.The Laboratory of Cell and Molecular BiologyNational Institutes of Diabetes and Digestive and Kidney Diseases, National Institutes of HealthBethesdaUSA
  3. 3.9000 Rockville Pike, Building 8 Room 319BethesdaUSA

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