Abstract
We describe a protocol for transient gene expression in rice protoplasts and its application to the study of Magnaporthe oryzae avirulence (AVR) gene function. In this assay the gene encoding the firefly luciferase protein is transfected into rice protoplasts by electroporation together with the candidate AVR genes. The luminescence can then be used to assess the viability of rice protoplasts. The hypersensitive response (HR) caused by the interaction between M. oryzae AVR and rice R genes can subsequently be monitored by recording the decrease in luminescence from the transfected cells.
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References
Bart R, Chern M, Park C-J, Bartley L, Ronald PC (2006) A novel system for gene silencing using siRNAs in rice leaf and stem-derived protoplasts. Plant Methods 2:13
Chen S, Tao L, Zeng L, Vega-Sanchez ME, Umemura K, Wang G-L (2006) A highly efficient transient protoplast system for analyzing defence gene expression and protein-protein interactions in rice. Mol Plant Pathol 7:417–427
Yoshida K, Saitoh H, Fujisawa S, Kanzaki H, Matsumura H, Yoshida K, Tosa Y, Chuma I, Takano Y, Win J, Kamoun S, Terauchi R (2009) Association genetics reveals three novel avirulence genes from rice blast fungal pathogen Magnaporthe oryzae. Plant Cell 21: 1573–1591
Okuyama Y, Kanzaki H, Abe A, Yoshida K, Tamiru M, Saitoh H, Fujibe T, Matsumura H, Shenton M, Galam DC, Undan J, Ito A, Sone T, Terauchi R (2011) A multifaceted genomics approach allows the isolation of the rice Pia-blast resistance gene consisting of two adjacent NBS-LRR protein genes. Plant J 66:467–479
Kanzaki H, Yoshida K, Saitoh H, Fujisaki K, Hirabuchi A, Alaux L, Fournier E, Tharreau D, Terauchi R (2012) Arms race co-evolution of Magnaporthe oryzae AVR-Pik and rice Pik genes driven by their physical interactions. Plant J 72:894–907
Acknowledgements
This work was supported by the Ministry of Agriculture, Forestry, and Fisheries of Japan (Genomics for Agricultural Innovation PMI-0010) and the Program for Promotion of Basic Research Activities for Innovative Biosciences (PROBRAIN), Japan, and Grant-in-aid for Scientific Research from the Ministry of Education, Cultures, Sports and Technology, Japan to HS and RT (Grant-in-Aid for Scientific Research on Innovative Areas 23113009).
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Kanzaki, H., Yoshida, K., Saitoh, H., Tamiru, M., Terauchi, R. (2014). Protoplast Cell Death Assay to Study Magnaporthe oryzae AVR Gene Function in Rice. In: Birch, P., Jones, J., Bos, J. (eds) Plant-Pathogen Interactions. Methods in Molecular Biology, vol 1127. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-986-4_20
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DOI: https://doi.org/10.1007/978-1-62703-986-4_20
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Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-1-62703-985-7
Online ISBN: 978-1-62703-986-4
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