Potato virus Y (PVY) induces the formation of lesions on infected leaves of potato plants cv. Rywal as a part of defense mechanism, called hypersensitive response conferred resistance. In this master thesis, we followed stromule formation and changes in chloroplastic redox state in pt-roGFP expressing potato plants after infection with PVY using confocal microscope and image analysis script for detection of redox state. We compared the response of genetically modified potato plants with reduced accumulation of apoplastic ROS (apoROS), chloroplastic ROS (chlROS) as well as varying levels of salicylic acid (SA) in different combinations. Carrying a detailed spatiotemporal analysis, we revealed that that the most oxidized chloroplasts accompanied by higher stromule count were predominantly located in the proximity of lesions. Furthermore, we confirmed the pivotal role of SA and chlROS in regulation of stromule formation, as increased SA levels induce it, while reduced SA levels and chlROS accumulation inhibit it. In order to asses the applicability and stability of the newly developed PVY-mTurquoise2 viral clone, we infected tobacco plants (Nicotina clevelandii) as well as genetically modified potato plants, described above. We confirmed that the dynamics of PVY-mTurquoise2 spread closely resembles the one of PVY-GFP and that PVY-mTurquoise remains stable in tobacco plants after several passages. Moreover, we demonstrated the precise identification of PVY-mTurquoise2 infected cells in inoculated potato plants using a confocal microscope, which allowed us to investigate stromule formation and chloroplastic redox state in immediate vicinity of those cells. Additionally, we presented an effective and optimized method for tracking PVY-GFP and PVY-mTurquoise2 viral clones using a luminometer.
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