The full-length sequence of AtFDH1 with native promoter was cloned into pMDC107 for GFP expression (AtFDH1-GFP). Stable Arabidopsis transgenic lines for the expression of AtFDH1-GFP were developed by floral dip transformation [48 (link)]. The localization of AtFDH1-GFP in epidermal cells was determined under the confocal laser scanning microscope (NIKON, Japan).
To observe the localization of AtFDH1, Arabidopsis wild-type Col-0 and AtFDH1-GFP expressing (under the control of AtFDH1 promoter) transgenic plants in Col-0 were grown in ½ MS media for four weeks, and AtFDH1-GFP expression in epidermal cells of Arabidopsis was visualized using a confocal laser scanning microscope (NIKON, Japan). The leaf tissues were floated with the bacterial suspension of host pathogen P. syringae pv. maculicola (1×106 CFU/ml) and nonhost pathogen P. syringae pv. tabaci (1×106 CFU/ml). After one hour inoculation, the leaf tissues were washed with distilled water, and localization of FDH1-GFP was observed. For wounding stress, the adaxial epidermal peels from wild-type Col-0 and AtFDH1-GFP expressing transgenic plants were prepared in the MES buffer (10 mM, pH 6.5), and localization of AtFDH1 was imaged under the confocal laser scanning microscope (NIKON, Japan).
To observe the localization of AtFDH1, Arabidopsis wild-type Col-0 and AtFDH1-GFP expressing (under the control of AtFDH1 promoter) transgenic plants in Col-0 were grown in ½ MS media for four weeks, and AtFDH1-GFP expression in epidermal cells of Arabidopsis was visualized using a confocal laser scanning microscope (NIKON, Japan). The leaf tissues were floated with the bacterial suspension of host pathogen P. syringae pv. maculicola (1×106 CFU/ml) and nonhost pathogen P. syringae pv. tabaci (1×106 CFU/ml). After one hour inoculation, the leaf tissues were washed with distilled water, and localization of FDH1-GFP was observed. For wounding stress, the adaxial epidermal peels from wild-type Col-0 and AtFDH1-GFP expressing transgenic plants were prepared in the MES buffer (10 mM, pH 6.5), and localization of AtFDH1 was imaged under the confocal laser scanning microscope (NIKON, Japan).
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