Zen blue lite program

For imaging, leaf sections approximately 1 cm 2 were excised, and at least four different leaves from at least three different plants were imaged for each construct. For leaf transient assays, N. tabacum was used for all Rips from K60; N. benthamiana was used for all Rips from GMI1000. For N. benthamiana, leaf sections were examined with confocal microscopy using a Nikon A1 spectral confocal microscope using staff support at the University of Minnesota Imaging Centers. Images were processed using the NIS Elements software package (Nikon, Tokyo, Japan). For N. tabacum, leaf sections were imaged using a Zeiss 880 LSM Upright Confocal microscope (Carl Zeiss Microscopy, White Plains, NY) at Purdue University. Leaf sections were examined on the abaxial side. GFP was excited at 488 nm and detected between 525 and 550 nm, and mCherry was excited at 561 nm and detected at 610 nm. Images captured on the Zeiss Confocal were processed using the Zeiss Zen Blue Lite program (Carl Zeiss). For plasmolysis experiments, samples were flooded with 1 M KNO 3 . After 5 min, leaf tissue was imaged. All images were one slice.

Live-cell imaging of N. benthamiana epidermal cells was performed 24 hours following agroinfiltration using a Zeiss LSM880 Axio Examiner upright confocal microscope as described previously (Denne et al., 2021) . Briefly, N. benthamiana leaf sections were excised and mounted in sterile water between a slide and a coverslip (adaxial surface toward the objective) and subsequently imaged using a Plan Apochromat 20x/0.8 objective, pinhole 1.0 AU. For plasmolysis, leaf sections were prepared as described above, submerged in 0.8 M mannitol solution for 20 minutes, and imaged shortly thereafter. The sYFP protein fusions were excited using a 514-nm argon laser and fluorescence was detected between 517-562 nm. Fluorescence from the and is also made available for use under a CC0 license.
was not certified by peer review) is the author/funder. This article is a US Government work. It is not subject to copyright under 17 USC 105
The copyright holder for this preprint (which this version posted May 25, 2022. ; https://doi.org/10.1101/2022.05.24.492667 doi: bioRxiv preprint Helm and Singh, et al 11 mCherry-tagged constructs was excited with a 561-nm helium-neon laser and detected between 565-669 nm. All confocal micrographs were captured on the Zeiss LSM880 upright confocal microscope and processed using the Zeiss Zen Blue Lite program (Carl Zeiss Microscopy, USA).