Evaluation of PTD-GFP fluorescence by confocal microscope in tobacco and lettuce leaf discs after <i>E. carotovora</i> infection.

<p>Leaf discs (9 mm in diameter) made using cork borer were infected with <i>E. carotovora</i> (OD₆₀₀ = 0.2) in a multi-well culture plate and plant cells were imaged by confocal microscopy. Representative images are from 67 stored images out of 189 observations of cells near the periphery of tobacco leaf discs incubated with <i>E. carotovora</i> for 30 min (A) or 1 hr (B). Different stages of GFP release were indicated by arrows and numbers. Arrow #1 represents very early stage of GFP release, showing negligible loss of GFP fluorescence. Arrow #2 represents intermediate step of GFP release demonstrating both GFP and chlorophyll fluorescence in chloroplasts. Arrow #3 represents late stage of GFP release outside of chloroplasts with detection of only red chlorophyll fluorescence. (C) Representative image from 70 stored images out of 194 observations of leaf discs of tobacco without <i>E. carotovora</i> inoculation. (D) Representative images are from 74 stored images out of 191 observations of cells near the periphery of lettuce leaf discs incubated with <i>E. carotovora</i> for 1 hr. (E) Representative images are from 68 stored images out of 153 observations of lettuce leaf discs without <i>E. carotovora</i> inoculation. Each experiment was repeated five times and each time two leaves from at least two different plants were used to make 6 discs per treatment. Bars represent 10 µm.