Abstract
Fluorescent proteins (FPs) remarkably advanced the study of cellular biology of plants. The most common application is their use as reporter proteins to determine the subcellular localization of a protein of interest (POI) by endogenous expression of a suitable FP-POI fusion construct in plant cells. In this chapter we describe three approaches, namely, particle bombardment, protoplast transformation, and Agrobacterium infiltration, to transiently express such fusion constructs in plant cells of different species. These approaches are versatile and can be utilized for diverse fluorescent protein-based applications.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Dixit R, Cyr R, Gilroy S (2006) Using intrinsically fluorescent proteins for plant cell imaging. Plant J 45(4):599–615
Berg RH, Beachy RN (2008) Fluorescent protein applications in plants. Methods Cell Biol 85:153–177
Xing S, Wallmeroth N, Berendzen KW, Grefen C (2016) Techniques for the analysis of protein-protein interactions in vivo. Plant Physiol 171(2):727–758
Grossmann G, Krebs M, Maizel A, Stahl Y, Vermeer JE, Ott T (2018) Green light for quantitative live-cell imaging in plants. J Cell Sci 131(2):jcs209270
Klein TM, Arentzen R, Lewis PA, Fitzpatrick-McElligott S (1992) Transformation of microbes, plants and animals by particle bombardment. Biotechnology 10(3):286–291
Abel S, Theologis A (1994) Transient transformation of Arabidopsis leaf protoplasts: a versatile experimental system to study gene expression. Plant J 5(3):421–427
Wroblewski T, Tomczak A, Michelmore R (2005) Optimization of Agrobacterium-mediated transient assays of gene expression in lettuce, tomato and Arabidopsis. Plant Biotechnol J 3(2):259–273
Sharma M, Bennewitz B, Klösgen RB (2018) Rather rule than exception? How to evaluate the relevance of dual protein targeting to mitochondria and chloroplasts. Photosynth Res 138(3):335–343
Rödiger A, Baudisch B, Langner U, Klösgen RB (2011) Dual targeting of a mitochondrial protein: the case study of cytochrome c1. Mol Plant 4(4):679–687
Wu FH, Shen SC, Lee LY, Lee SH, Chan MT, Lin CS (2009) Tape-Arabidopsis Sandwich-a simpler Arabidopsis protoplast isolation method. Plant Methods 5(1):1–10
Sharma M, Bennewitz B, Klösgen RB (2018) Dual or not dual? – Comparative analysis of fluorescence microscopy-based approaches to study organelle targeting specificity of nuclear-encoded plant proteins. Front Plant Sci 1350
Snapp E (2005) Design and use of fluorescent fusion proteins in cell biology. Curr Protoc Cell Biol 27(1):21–24
Hooper CM, Castleden IR, Tanz SK, Aryamanesh N, Millar AH (2017) SUBA4: the interactive data analysis centre for Arabidopsis subcellular protein locations. Nucleic Acids Res 45(D1):D1064–D1074
Baudisch B, Langner U, Garz I, Klösgen RB (2014) The exception proves the rule? Dual targeting of nuclear-encoded proteins into endosymbiotic organelles. New Phytol 201(1):80–90
Komari T, Takakura Y, Ueki J, Kato N, Ishida Y, Hiei Y (2006) Binary vectors and super-binary vectors. Agrobacterium Protoc:15–42
Überlacker B, Werr W (1996) Vectors with rare-cutter restriction enzyme sites for expression of open reading frames in transgenic plants. Mol Breed 2(3):293–295
Lee S, Su G, Lasserre E, Aghazadeh MA, Murai N (2012) Small high-yielding binary Ti vectors pLSU with co-directional replicons for Agrobacterium tumefaciens-mediated transformation of higher plants. Plant Sci 187:49–58
Sharma M (2019) Determining targeting specificity of nuclear-encoded organelle proteins. Dissertation, Martin-Luther-Universität Halle-Wittenberg, Germany. https://doi.org/10.25673/13979
Acknowledgments
MS was supported by a fellowship from the BRAVE project funded by the ERASMUS MUNDUS Action 2 program of the European Union.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Sharma, M., Klösgen, R.B., Bennewitz, B. (2023). Fluorescent Protein-Based Approaches for Subcellular Protein Localization in Plants. In: Sharma, M. (eds) Fluorescent Proteins. Methods in Molecular Biology, vol 2564. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2667-2_9
Download citation
DOI: https://doi.org/10.1007/978-1-0716-2667-2_9
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-2666-5
Online ISBN: 978-1-0716-2667-2
eBook Packages: Springer Protocols