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Elife. 2020 Feb 11;9. pii: e52546. doi: 10.7554/eLife.52546.

Imaging plant germline differentiation within Arabidopsis flowers by light sheet microscopy.

Author information

1
Central European Institute of Technology (CEITEC), Masaryk University, Brno, Czech Republic.
2
IT4Innovations, VSB-Technical University of Ostrava, Ostrava, Czech Republic.
3
Department of Computer Science, FEECS VSB - Technical University of Ostrava, Ostrava, Czech Republic.
4
Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Seeland, Germany.
5
Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany.
#
Contributed equally

Abstract

In higher plants, germline differentiation occurs during a relatively short period within developing flowers. Understanding of the mechanisms that govern germline differentiation lags behind other plant developmental processes. This is largely because the germline is restricted to relatively few cells buried deep within floral tissues, which makes them difficult to study. To overcome this limitation, we have developed a methodology for live imaging of the germ cell lineage within floral organs of Arabidopsis using light sheet fluorescence microscopy. We have established reporter lines, cultivation conditions, and imaging protocols for high-resolution microscopy of developing flowers continuously for up to several days. We used multiview imagining to reconstruct a three-dimensional model of a flower at subcellular resolution. We demonstrate the power of this approach by capturing male and female meiosis, asymmetric pollen division, movement of meiotic chromosomes, and unusual restitution mitosis in tapetum cells. This method will enable new avenues of research into plant sexual reproduction.

KEYWORDS:

A. thaliana; SPIM; cell biology; flower; germline; light sheet microscopy; live cell imaging; meiosis; plant biology

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