We describe a fluorescence imaging method to visualize the dynamics of the central vacuole in rice cells during invasion by the blast fungus Magnaporthe oryzae. This method utilizes the combination of confocal microscopy, rice sheath cells (optically transparent), fluorescently tagged M. oryzae (red fluorescence), and fluorescein diacetate staining (green fluorescence; visualizing vacuole dynamics). Using this method, we demonstrate that the vacuole undergoes progressive shrinkage and collapse during M. oryzae infection.
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We thank Dong Won Kim, Jie Zhu, and Mariel Pfeifer in the Khang Lab (http://www.khanglab.org/) for their help and discussions. We acknowledge the assistance of the Biomedical Microscopy Core at the University of Georgia with imaging using a Zeiss LSM 510 confocal microscope. This work was supported by the Agriculture and Food Research Initiative competitive grants program, Award number 2014-67013-21717 from the USDA National Institute of Food and Agriculture.
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