Skip to main content

Studying Arabidopsis Chloroplast Structural Organisation Using Transmission Electron Microscopy

  • Protocol
  • First Online:
Chloroplast Research in Arabidopsis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 774))

Abstract

Chloroplasts, as well as other, non-photosynthetic types of plastid, are characteristic structures within plant cells. They are relatively large organelles (typically 1–5 μm in diameter), and so can readily be analysed by electron microscopy. Chloroplast structure is remarkably complex, comprising at least six distinct sub-organellar compartments, and is sensitive to developmental changes, environmental effects, and genetic lesions. Transmission electron microscopy (TEM), therefore, represents a powerful technique for monitoring the effects of various changing parameters or treatments on the development and differentiation of these important organelles. We describe a method for the analysis of Arabidopsis plant material by TEM, primarily for the assessment of plastid ultrastructure.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 159.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Whatley, J. M. (1978) A suggested cycle of plastid developmental interrelationships. New Phytol. 80, 489–502.

    Article  Google Scholar 

  2. López-Juez, E., and Pyke, K. A. (2005) Plastids unleashed: their development and their integration in plant development. Int. J. Dev. Biol. 49, 557–577.

    Article  PubMed  Google Scholar 

  3. Kinsman, E. A., and Pyke, K. A. (1998) Bundle sheath cells and cell-specific plastid development in Arabidopsis leaves. Development 125, 1815–1822.

    PubMed  CAS  Google Scholar 

  4. Schelbert, S., Aubry, S., Burla, B., Agne, B., Kessler, F., Krupinska, K., and Hörtensteiner, S. (2009) Pheophytin pheophorbide hydrolase (pheophytinase) is involved in chlorophyll breakdown during leaf senescence in Arabidopsis. Plant Cell 21, 767–785.

    Article  PubMed  CAS  Google Scholar 

  5. Kubis, S., Patel, R., Combe, J., Bédard, J., Kovacheva, S., Lilley, K., Biehl, A., Leister, D., Ríos, G., Koncz, C., and Jarvis, P. (2004) Functional specialization amongst the Arabidopsis Toc159 family of chloroplast protein import receptors. Plant Cell 16, 2059–2077.

    Article  PubMed  CAS  Google Scholar 

  6. Kuo, J. (2007) Electron Microscopy: Methods and Protocols. Methods in Molecular Biology, Vol. 369. Humana Press, Totowa, NJ, USA.

    Google Scholar 

  7. Pfeiffer, S., and Krupinska, K. (2005) Chloroplast ultrastructure in leaves of Urtica dioica L. analyzed after high-pressure freezing and freeze-substitution and compared with conventional fixation followed by room temperature dehydration. Microsc. Res. Tech. 68, 368–376.

    Article  PubMed  Google Scholar 

  8. Hayat, M. A. (1981) Fixation for Electron Microscopy. Academic Press, New York, USA.

    Google Scholar 

  9. Glauert, A. M., and Lewis, P. R. (1998) Biological Specimen Preparation for Transmission Electron Microscopy. Practical Methods in Electron Microscopy, Vol. 17. Portland Press Ltd., London, UK.

    Google Scholar 

  10. Li, H., Culligan, K., Dixon, R. A., and Chory, J. (1995) CUE1: a mesophyll cell-specific positive regulator of light-controlled gene expression in Arabidopsis. Plant Cell 7, 1599–1610.

    Article  PubMed  CAS  Google Scholar 

  11. Ellis, E. A. (2006) Solutions to the problem of substitution of ERL 4221 for vinyl cyclohexene dioxide in Spurr low viscosity embedding formulations. Microscopy Today 14, 32–33.

    CAS  Google Scholar 

  12. Reynolds, E. S. (1963) The use of lead citrate at high pH as an electron-opaque stain in electron microscopy. J. Cell Biol. 17, 208–212.

    Article  PubMed  CAS  Google Scholar 

  13. Ellis, E. A. (2007) Poststaining grids for transmission electron microscopy: conventional and alternative protocols. In, Electron Microscopy: Methods and Protocols (Kuo, J., ed.) Humana Press, Totowa, NJ, USA, pp. 97–106.

    Google Scholar 

  14. Hunter, E. (1993) Practical Electron Microscopy: A Beginner’s Illustrated Guide. Cambridge University Press, Cambridge, UK.

    Google Scholar 

  15. Kovacheva, S., Bédard, J., Patel, R., Dudley, P., Twell, D., Ríos, G., Koncz, C., and Jarvis, P. (2005) In vivo studies on the roles of Tic110, Tic40 and Hsp93 during chloroplast protein import. Plant J. 41, 412–428.

    Article  PubMed  CAS  Google Scholar 

  16. Aronsson, H., Boij, P., Patel, R., Wardle, A., Töpel, M., and Jarvis, P. (2007) Toc64/OEP64 is not essential for the efficient import of proteins into chloroplasts in Arabidopsis thaliana. Plant J. 52, 53–68.

    Article  PubMed  CAS  Google Scholar 

  17. Hall, J. L., and Hawes, C. (1991) Electron Microscopy of Plant Cells. Academic Press, London, UK.

    Google Scholar 

  18. Bozzola, J. J. (2007) Conventional specimen preparation techniques for transmission electron microscopy of cultured cells. In, Electron Microscopy: Methods and Protocols (Kuo, J., ed.) Humana Press, Totowa, NJ, USA, pp. 1–18.

    Google Scholar 

  19. Spurr, A. R. (1969) A low-viscosity epoxy resin embedding medium for electron microscopy. J. Ultrastruct. Res. 26, 31–43.

    Article  PubMed  CAS  Google Scholar 

  20. Kuo, J. (2007) Processing plant tissues for ultrastructural study. In, Electron Microscopy: Methods and Protocols (Kuo, J., ed.) Humana Press, Totowa, NJ, USA, pp. 35–45.

    Google Scholar 

  21. Louw, J., Williams, K., Harper, I. S., and Walfe-Coote, S. A. (1990) Electron dense artefactual deposits in tissue sections: the role of ethanol, uranyl acetate and phosphate buffer. Stain Technol. 65, 243–250.

    PubMed  CAS  Google Scholar 

Download references

Acknowledgments

The authors wish to acknowledge Ms. Natalie Allcock of the Core Biotechnology Services Electron Microscopy Laboratory, University of Leicester, for both technical support and comments on the manuscript.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Stefan Hyman .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2011 Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Hyman, S., Jarvis, R.P. (2011). Studying Arabidopsis Chloroplast Structural Organisation Using Transmission Electron Microscopy. In: Jarvis, R. (eds) Chloroplast Research in Arabidopsis. Methods in Molecular Biology, vol 774. Humana Press. https://doi.org/10.1007/978-1-61779-234-2_8

Download citation

  • DOI: https://doi.org/10.1007/978-1-61779-234-2_8

  • Published:

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-61779-233-5

  • Online ISBN: 978-1-61779-234-2

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics