Shanghai Key Laboratory of Plant Molecular Sciences, College of Life Sciences, Shanghai Normal University, Shanghai, 200234, China.
Plant Reprod. 2021 Jun;34(2):91-101. doi: 10.1007/s00497-021-00412-5. Epub 2021 Apr 26.
The pollen coat, which forms on the pollen surface, consists of a lipid-protein matrix. It protects pollen from desiccation and is involved in adhesion, pollen-stigma recognition, and pollen hydration during interactions with the stigma. The classical methods used for pollen coat observation are scanning and transmission electron microscopy. In this work, we screened a collection of fluorescence dyes and identified two fluorescent brighteners FB-52 and FB-184. When they were used together with the exine-specific dye, Basic fuchsin, the pollen coat and the exine structures could be clearly visualized in the pollen of Brassica napus. This co-staining method was applied successfully in staining pollen from Fraxinus chinensis, Calystegia hederacea, and Petunia hybrida. Using this method, small pollen coat-containing cavities were detected in the outer pollen wall layer of Oryza sativa and Zea mays. We further showed these dyes are compatible with fluorescent protein markers. In the Arabidopsis thaliana transgenic line of GFP-tagged pollen coat protein GRP19, GRP19-GFP was observed to form particles at the periphery of pollen coat. This simple staining method is expected to be widely used for the studies of the palynology as well as the pollen-stigma interaction.
花粉外壁在花粉表面形成，由脂-蛋白基质组成。它保护花粉免受干燥，并参与与柱头的相互作用过程中的粘附、花粉-柱头识别和花粉水合作用。用于观察花粉外壁的经典方法是扫描和透射电子显微镜。在这项工作中，我们筛选了一系列荧光染料，并鉴定出两种荧光增白剂 FB-52 和 FB-184。当它们与外壁特异性染料碱性品红一起使用时，油菜花粉的花粉外壁和外壁结构可以清晰地观察到。这种共染色方法成功地应用于染色荆芥、旋花和矮牵牛的花粉。使用这种方法，在水稻和玉米的花粉外壁层中检测到了含有小花粉外壁的腔。我们进一步表明，这些染料与荧光蛋白标记物兼容。在 GFP 标记花粉外壁蛋白 GRP19 的拟南芥转基因系中，观察到 GRP19-GFP 在花粉外壁的周边形成颗粒。这种简单的染色方法有望广泛应用于孢粉学研究以及花粉-柱头相互作用的研究。
