Journal: Scientific Reports

Article Title: Phytocyanin-encoding genes confer enhanced ozone tolerance in Arabidopsis thaliana

doi: 10.1038/s41598-022-25706-0

Figure Lengend Snippet: Cellular localisation and the ability of AtUC5-GFP fusion proteins to confer O 3 tolerance. ( a ) Diagrams depicting the chimeric genes to produce fusion proteins in transgenic plants. ( b ) Results of western blotting to detect fusion protein expression in extracts of transgenic plants. Equal amounts (3 µg) of protein extracted from plants of various lines were subjected to electrophoresis and probed with anti-GFP antibody using Simple Western™ (Bio-Techne, Minneapolis, USA). GFP was added to the protein extract from the wild-type plant just before the electrophoresis for the lane “wt + GFP.” Results were obtained as digital data for each sample and exhibited as band images as their full-length blots. ( c ) Microscopic observation of GFP fluorescence in epidermal tissues of transgenic plants. α fluorescence images of epidermal tissue of a leaf transgenic for 35S:GFP-AtUC5 . β fluorescence image of epidermal tissue of a leaf transgenic for 35S:AtUC5-GFP . γ fluorescence image of epidermal tissue of a leaf transgenic for 35S:GFP-AtUC5 after treatment with hyperosmotic solution. δ fluorescence image of the epidermal tissue of a leaf transgenic for 35S:AtUC5-GFP after treatment with hyperosmotic solution. Arrowheads and arrows denote stomata and apoplastic matrices, respectively. ( d ) Diagrams depicting the presumed structures and localizations of fusion proteins in transgenic plants. In 35S:GFP-AtUC5 plants, the fusion protein is released from the plasma membrane after digestion at the GPI moiety by phospholipases. ( e ) Images of various lines 1 day after 2 h exposure to fresh air or 0.3 µL L −1 O 3 under 420 µmol photons m −2 s −1 illumination. Wt wild-type plants, GFP green fluorescent protein, 35S:GFP-AtUC5 35S:AtUC5-GFP , transgenic plants producing fusion proteins, 35S cauliflower mosaic virus 35S promoter, N a fragment encoding the N-terminal signal peptide of AtUC5, PLD plastocyanin-like domain, ALR arabinogalactan protein-like region, GPI glycosylphosphatidylinositol moiety, CGAS C-terminal glycosylphosphatidylinositol-anchored signal, FA fresh air .

Article Snippet: Equal amounts (3 µg) of protein extracted from plants of various lines were subjected to electrophoresis and probed with anti-GFP antibody using Simple Western™ (Bio-Techne, Minneapolis, USA).

Techniques: Transgenic Assay, Western Blot, Expressing, Electrophoresis, Fluorescence, Membrane, Virus