We describe a simple method to view meiotic cells in whole anthers from a range of plants. The method retains spatial organisation and enables simultaneous analysis of many meiotic cells.
Understanding the process of male meiosis in flowering plants, and the role of genes involved in this process, offers potential for plant breeding, such as through increasing the level of genetic variation or the manipulation of ploidy levels in the gametes. A key to the characterisation of meiotic gene function and meiosis in non-model crop plants, is the analysis of cells undergoing meiosis, a task made difficult by the inaccessible nature of these cells. Here, we describe a simple and rapid method to analyse plant male meiosis in intact anthers in a range of plant species. This method allows analysis of numerous cells undergoing meiosis and, as meiotic cells stay within the anther, it retains information of the three-dimensional organisation and the location of organelles in meiotic cells. We show that the technique provides information on male meiosis by looking at the synchrony of meiotic progression between and within locules, and comparing wildtype and mutant plants through the chromosome separation stages in Arabidopsis thaliana. Additionally, we demonstrate that the protocol can be adopted to other plants with different floral morphology using Medicago truncatula as an example with small floral buds and the non-model plant kiwifruit (Actinidia chinensis) with larger buds and anthers.
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We would like to thank Shona Seymour at Plant & Food Research for the collection of kiwifruit budwood. This work was funded by the Department of Biochemistry, University of Otago, New Zealand and the Endeavour Fund from the Ministry of Business Innovation and Employment (Smart Idea UOOX1801).
Department of Biochemistry, University of Otago and Ministry of Business Innovation and Employment Endeavour Fund (Smart Idea UOOX1801).
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Communicated by Dolf Weijers.
Below is the link to the Supplementary Information.
Single optical section from CLSM analysis of an anther from A. thaliana Ler. The anther was isolated from a flower bud, chromosomal and organelle DNA stained with DAPI and observed with CLSM. A z-series for this anther is shown in Online Resource 2. Two locules are visible, one with meiocytes of metaphase II (MII) and one at telophase II (TII). Not all nuclei are visible in the optical section. An arrow indicates a binucleate tapetal cell and an arrowhead indicates an organelle band between two chromosome groups in meiosis II. Scale, 50 µm (TIF 5538 kb)
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Rossig, C., Le Lievre, L., Pilkington, S.M. et al. A simple and rapid method for imaging male meiotic cells in anthers of model and non-model plant species. Plant Reprod 34, 37–46 (2021). https://doi.org/10.1007/s00497-021-00404-5
- Male meiosis
- Confocal laser scanning microscopy
- Fluorescent staining