Live imaging was performed as previously described (Valuchova et al., 2020 (link)). Floral buds .3 to .7 mm long were selected from the main inflorescence. The reproductive organs were exposed by removing the sepals and placed into glass capillaries (size 4, Zeiss) containing 1/2 Murashige and Skoog (MS) medium (5% sucrose, pH 5.8) with 1% low melting point agarose (Sigma Aldrich). The inflorescence embedded in solidified MS was pushed out of the capillary and then placed into the capillary holder for the Z.1 ZEISS light‐sheet microscope. After the holder is inserted in the microscope chamber, the remaining space is filled with liquid 1/2 MS medium (5% sucrose, pH 5.8). Images were taken every 5 min with a 10× objective (detection optics 10×/.5), single illumination (illumination optics 10×/.2), 561‐nm laser (15% intensity). The large raw data files were processed by ZEN Blue software (Zeiss).
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Live Imaging of Floral Bud Development
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Agarose
Capillary
Illumination
Inflorescence
Light microscope
Microscope
Optics
Reproductive organs
Sucrose
Corresponding Organization :
Other organizations : Central European Institute of Technology, Central European Institute of Technology – Masaryk University, Masaryk University, Gregor Mendel Institute of Molecular Plant Biology, Austrian Academy of Sciences, Universität Hamburg
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Variable analysis
independent variables
- Floral bud size (.3 to .7 mm long)
- Reproductive organ development
- Removing the sepals to expose the reproductive organs
- Glass capillaries (size 4, Zeiss) containing 1/2 Murashige and Skoog (MS) medium (5% sucrose, pH 5.8) with 1% low melting point agarose (Sigma Aldrich)
- Illumination (10×/.2 illumination optics), 561‐nm laser (15% intensity)
- Imaging with a 10× objective (detection optics 10×/.5)
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