Suppression of necrotic effects by foliar spray application of high concentration of AsA. A, Syringe agroinfiltration was performed to express GFP, hCul1, and hFbxw7 in the leaves of N. benthamiana, followed by foliar spray application of AsA at the indicated concentration at 2-d intervals. The plants were incubated at 20°C for 7 d. Soluble proteins from the indicated fresh mass (FM) of these leaves were separated by SDS-PAGE. GFP (B) or hCul1 (D) were detected by Coomassie Brilliant Blue (CBB) staining or anti-DYKDDDDK antibody. NT indicates nontransfected plants. The amount of GFP (C) or hCul1 (E) was determined from band intensities. N. benthamiana plants agroinfiltrated with different vectors were incubated at 25°C following the foliar spray application of 200 mM AsA, and soluble proteins were extracted from leaves at 3, 5, or 7 d after agroinfiltration. hCul1 (F) or hFbxw7 (H) were detected with anti-DYKDDDDK antibody. The amount of hCul1 (G) were determined from band intensities. Data represent the means ± sd (n = 3–4) and significance was determined using unpaired Student’s t tests (*P < 0.05) (C, E, G).