Supplemental Movie 1.
Time-lapse fluorescence imaging of germinating Arabidopsis pollen using Spinning Disk Confocal Microscopy. Pollen expressing GFP-tagged Armadillo Repeat Only 1 (ARO1) was germinated in vitro in a self-made multi-well micro-germination setup on solidified pollen germination medium (PGM) containing 10 µM 24-Epibrassinolide, a phytosterol from the brassinosteroid family, to promote pollen germination and elongation, as described by Vogler et al. (Plant Reprod (2014) 27:153-167; DOI 10.1007/s00497-014-0247-x). Simultaneous imaging in an exemplary 10 well micro-germination setup was performed over 6 hours. Every position shows maximum intensity projections of 11 optical slices, imaged every 3 minutes. Pollen tubes start to emerge from the pollen grains 1h after incubation. Mounting pollen in this micro-germination setup allows simultaneous imaging of up to twelve (or even more) independent PGM pads under one cover slip. The inexpensive and easy-to-perform technique is amenable to every standard microscope (upright or inverse) and illumination technique (Epifluorescence, CLSM, Spinning Disc, TIRF, 2-Photon).
Time stamper (upper left) shows hours and minutes. Position B1 was readjusted after 1 h PT growth. Image processing was performed using the open source software ImageJ. Scale bar (lower right): 50 µm.