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Fig. 5 | Plant Methods

Fig. 5

From: Bioorthogonal click chemistry for fluorescence imaging of choline phospholipids in plants

Fig. 5

Dual visualization of choline phospholipids with nuclei, cell walls, and chloroplasts. Arabidopsis plants were grown for 6 weeks on agar media containing 200 µM propargylcholine (pp-Cho). a–c Roots were harvested and fixed, and fluorescein azide reactions were performed. Confocal laser scanning images show a, b root epidermal cells with fluorescein azide (green) marking propargyl-PC and a′, b′ DAPI fluorescence marking nuclei (blue). a″, b″ Merged images combine the fluorescence signals. Propargyl-PC signals typically surround (red arrow in b″) but do not overlay with nuclei. c Root epidermal cells were imaged after staining with propidium iodide (PI) to mark cell walls. c Fluorescein azide (green) indicates propargyl-PC and c′ PI marks cell walls. c″ Merged images show that the propargyl-PC occurs along cell walls (white arrow) but does not overlap with the cell wall stain. d, e Leaf samples were harvested and fluorescein azide reactions were performed followed by confocal microscopy. d, e Fluorescein azide (green) and d′, e′ chlorophyll fluorescence (red). d″, e″ Light microscopy. d‴, e‴ Merged images combine light microscopy overlaid with the fluorescence signals. Bright propargyl-PC foci appear independent of chlorophyll fluorescence. d Leaf mesodermal cells were imaged after epidermal layer was removed by peeling. A high density of chloroplasts (red) is visible. e Cross section showing fewer chloroplasts. Cell peripheries are marked by fluorescence signals from propargyl-PC. Co-localization was quantified using the Pearson correlation coefficient (PCC) (see Additional file 3: Table S1)

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