Abstract
Optogenetic tools to control gene expression have many advantages over the classical chemically inducible systems, overcoming intrinsic limitations of chemical inducers such as solubility, diffusion, and cell toxicity. They offer an unmatched spatiotemporal resolution and permit quantitative and noninvasive control of the gene expression. Here we describe a protocol of a synthetic light-inducible system for the targeted control of gene expression in plants based on the plant photoreceptor phytochrome B and one of its interacting factors (PIF6). The synthetic toggle switch system is in the ON state when plant protoplasts are illuminated with red light (660 nm) and can be returned to the OFF state by subsequent illumination with far-red light (760 nm). In this protocol, the implementation of a red light-inducible expression system in plants using Light-Emitting Diode (LED) illumination boxes is described, including the isolation and transient transformation of plant protoplasts from Arabidopsis thaliana and Nicotiana tabacum.
The original version of this chapter was revised. The erratum to this chapter is available at: DOI 10.1007/978-1-4939-3512-3_28
An erratum to this chapter can be found at http://dx.doi.org/10.1007/978-1-4939-3512-3_28
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Corrado G, Karali M (2009) Inducible gene expression systems and plant biotechnology. Biotechnol Adv 27(6):733–743
Junker A, Junker B (2012) Synthetic gene networks in plant systems. In: Weber W, Fussenegger M (eds) Synthetic gene networks, vol 813, Methods in molecular biology. Humana, New York, pp 343–358
Padidam M (2003) Chemically regulated gene expression in plants. Curr Opin Plant Biol 6(2):169–177
Zhang K, Cui B (2015) Optogenetic control of intracellular signaling pathways. Trends Biotechnol 33(2):92–100
Müller K, Naumann S, Weber W, Zurbriggen MD (2015) Optogenetics for gene expression in mammalian cells. Biol Chem 396(2):145–152
Müller K, Siegel D, Rodriguez Jahnke F, Gerrer K, Wend S, Decker EL, Reski R, Weber W, Zurbriggen MD (2014) A red light-controlled synthetic gene expression switch for plant systems. Mol BioSyst 10(7):1679–1688
Dovzhenko A, Bergen U, Koop HU (1998) Thin-alginate-layer technique for protoplast culture of tobacco leaf protoplasts: shoot formation in less than two weeks. Protoplasma 204(1-2):114–118
Dovzhenko A, Dal Bosco C, Meurer J, Koop HU (2003) Efficient regeneration from cotyledon protoplasts in Arabidopsis thaliana. Protoplasma 222(1–2):107–111
Luo Y, Koop H-U (1997) Somatic embryogenesis in cultured immature zygotic embryos and leaf protoplasts of Arabidopsis thaliana ecotypes. Planta 202(3):387–396
Menczel L, Galiba G, Nagy F, Maliga P (1982) Effect of radiation dosage on efficiency of chloroplast transfer by protoplast fusion in nicotiana. Genetics 100(3):487–495
Müller K, Zurbriggen MD, Weber W (2014) Control of gene expression using a red- and far-red light-responsive bi-stable toggle switch. Nat Protoc 9(3):622–632
Müller K, Engesser R, Metzger S, Schulz S, Kämpf MM, Busacker M, Steinberg T, Tomakidi P, Ehrbar M, Nagy F, Timmer J, Zubriggen MD, Weber W (2013) A red/far-red light-responsive bi-stable toggle switch to control gene expression in mammalian cells. Nucleic Acids Res 41(7):e77
Koop H-U, Steinmüller K, Wagner H, Rößler C, Eibl C, Sacher L (1996) Integration of foreign sequences into the tobacco plastome via polyethylene glycol-mediated protoplast transformation. Planta 199(2):193–201
Wend S, Bosco CD, Kämpf MM, Ren F, Palme K, Weber W, Dovzhenko A, Zurbriggen MD (2013) A quantitative ratiometric sensor for time-resolved analysis of auxin dynamics. Sci Rep 3:2052
Acknowledgments
This work was supported in part by the Excellence Initiative of the German Federal and State Governments (EXC294-BIOSS, GSC-4 Spemann Graduate School (SGBM)) and the Alexander von Humbolt Foundation (research Grant no. 1141629). We thank Susanne Knall and Frauke Bartels-Burgahn for experimental assistance. We thank J. Schmidt, D. Schächtele and J. Meßmer (University of Freiburg) for designing and constructing the illumination boxes.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2016 Springer Science+Business Media New York
About this protocol
Cite this protocol
Ochoa-Fernandez, R. et al. (2016). Optogenetics in Plants: Red/Far-Red Light Control of Gene Expression. In: Kianianmomeni, A. (eds) Optogenetics. Methods in Molecular Biology, vol 1408. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3512-3_9
Download citation
DOI: https://doi.org/10.1007/978-1-4939-3512-3_9
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-3510-9
Online ISBN: 978-1-4939-3512-3
eBook Packages: Springer Protocols