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. 2022 Sep 10;191(1):515–527. doi: 10.1093/plphys/kiac423

Figure 2.

Figure 2

Histochemical analysis and localization of DEF8. A–C, GUS expression driven by the DEF8 promoter at the seedling stage. Histochemical assay of 7-day-old rice seedlings (A), root cross section (B) and zoom in from (B) in dashed box (C). Arrow heads indicate phloem. Bar = 1 cm in (A), 50 µm in (B), and 10 µm in (C). D, Root cross section of rice seedlings harboring the construct proDEF8::fDEF8-mRFP. P, phloem; X, xylem. Bar = 25 μm. E–H, Subcellular localization of DEF8. Onion epidermal cells were transiently transformed with mRFP (E), 35S::fDEF8-mRFP (fDEF8-mRFP, F), and 35S::tDEF8-mRFP (tDEF8-mRFP, H), and incubated in 30% sucrose to induce plasmolysis before confocal microscopy imaging, and protoplasts (Pro) are indicated by arrows. G, DEF8 localization in transgenic rice harboring proDEF8::fDEF8-mRFP. Epidermal cells of leaf sheath were incubated in 30% sucrose to induce plasmolysis. I, Detection of DEF8 by Western blotting. Rice seedlings from (G) were grown for 2 weeks in hydroponics. Proteins were extracted from leaf, xylem sap (X-Sap), or guttation fluid, respectively, and subjected to Western blotting assay using an anti-RFP antibody. J, Effect of BFA on DEF8 localization in root. Four-day-old transgenic Col-0 expressing 35S::fDEF8-GFP was treated with 50 µM BFA for 30 min before imaging. DMSO-treated root cells were used as control.