Skip to main content

Identifying Subcellular Protein Localization with Fluorescent Protein Fusions After Transient Expression in Onion Epidermal Cells

  • Protocol
  • First Online:
Plant Cell Morphogenesis

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1080))

Abstract

Most biochemical functions of plant cells are carried out by proteins which act at very specific places within these cells, for example, within different organelles. Identifying the subcellular localization of proteins is therefore a useful tool to narrow down the possible functions that a novel or unknown protein may carry out. The discovery of genetically encoded fluorescent markers has made it possible to tag specific proteins and visualize them in vivo under a variety of conditions. This chapter describes a simple method to use transient expression of such fluorescently tagged proteins in onion epidermal cells to determine their subcellular localization relative to known markers.

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 119.00
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. Chalfie M, Tu Y, Euskirchen G et al (1994) Green fluorescent protein as a marker for gene expression. Science 263:802–805

    Article  PubMed  CAS  Google Scholar 

  2. Matz MV, Fradkov AF, Labas YA et al (1999) Fluorescent proteins from nonbioluminescent Anthozoa species. Nat Biotechnol 17:969–973

    Article  PubMed  CAS  Google Scholar 

  3. Rizzo MA, Springer GH, Granada B et al (2004) An improved cyan fluorescent protein variant useful for FRET. Nat Biotechnol 22: 445–449

    Article  PubMed  CAS  Google Scholar 

  4. Shaner NC, Campbell RE, Steinbach PA et al (2004) Improved monomeric red, orange and yellow fluorescent proteins derived from Discosoma sp. red fluorescent protein. Nat Biotechnol 22:1567–1572

    Article  PubMed  CAS  Google Scholar 

  5. Geldner N, Dénervaud-Tendon V, Hyman DL et al (2009) Rapid, combinatorial analysis of membrane compartments in intact plants with a multicolor marker set. Plant J 59:169–178

    Article  PubMed  CAS  Google Scholar 

  6. Nelson BK, Cai X, Nebenführ A (2007) A multi-color set of in vivo organelle markers for colocalization studies in Arabidopsis and other plants. Plant J 51:1126–1136

    Article  PubMed  CAS  Google Scholar 

  7. Batoko H, Zheng H-Q, Hawes C et al (2000) A Rab1 GTPase is required for transport between the endoplasmic reticulum and Golgi apparatus and for normal Golgi movement in plants. Plant Cell 12:2201–2218

    PubMed  CAS  Google Scholar 

  8. Li J-F, Park E, von Arnim AG et al (2009) The FAST technique: a simplified Agrobacterium-based transformation method for transient gene expression analysis in seedlings of Arabidopsis and other plant species. Plant Methods 5:6

    Article  PubMed  Google Scholar 

  9. Marion J, Bach L, Bellec Y et al (2008) Systematic analysis of protein subcellular localization and interaction using high-throughput transient transformation of Arabidopsis seedlings. Plant J 56:169–179

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

I thank the members of my lab for numerous improvements of the procedure. Work in my lab is supported by the National Science Foundation (MCB-0822111).

Author information

Authors and Affiliations

Authors

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2014 Springer Science+Business Media, New York

About this protocol

Cite this protocol

Nebenführ, A. (2014). Identifying Subcellular Protein Localization with Fluorescent Protein Fusions After Transient Expression in Onion Epidermal Cells. In: Žárský, V., Cvrčková, F. (eds) Plant Cell Morphogenesis. Methods in Molecular Biology, vol 1080. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-643-6_6

Download citation

  • DOI: https://doi.org/10.1007/978-1-62703-643-6_6

  • Published:

  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-642-9

  • Online ISBN: 978-1-62703-643-6

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics