Figure 5

Direct leaf disc analysis of engineered monomeric fluorescent proteins. Fluorescent proteins were transiently expressed by co-infiltrating N. benthamiana leaves with an Agrobacterium pSN.5 p19 culture plus cultures of Agrobacterium containing pSN.5 mTagBFP2 (mTagBFP2), pSN.5 mTFP1 (mTFP1), pSN.5 mNeon (mNeon), pSN.5 mPapaya1 (mPapaya1), pSN.5 TagRFP-T (TagRFP-T) or a strain with no expression vector (Φ). (A) At 6 dpa, cell fluorescence was imaged by confocal microscopy. Fluorophore excitation (dotted lines) and emission spectra (solid lines) from agro-infiltrated leaf discs were measured in a 96-well plate reader. Leaves infiltrated with an Agrobacterium culture without expression vectors were used as control (black lines). Relative fluorescence intensity (RFI) was plotted using fluorophore peaks equal to 100. UV wavelengths are in gray, visible spectrum colors were assigned as described [39]. (B) Box-plot graphs show quantification values from n = 8 samples/condition. Fluorescence intensity of the leaf discs agro-infiltrated with the indicated fluorescent protein-expressing plasmid or without expression vector (Φ) was measured in a monochromator-based plate reader. RFI was plotted using each fluorophore mean value equal to 100.