SummaryThe isolation of male and female gametes is an effective method to study the fertilization mechanisms of higher plants. An osmotic shock method was used to rupture pollen grains of Allium tuberosum Roxb and release the pollen contents, including generative cells, which were mass collected. The pollinated styles were cut following 3 h of in vivo growth, and cultured in medium for 6–8 h, during which time pollen tubes grew out of the cut end of the style. After pollen tubes were transferred into a solution containing 6% mannitol, tubes burst and released pairs of sperm cells. Ovules of A. tuberosum were incubated in an enzyme solution for 30 min, and then dissected to remove the integuments. Following transfer to a dissecting solution free of enzymes, each nucellus was cut in the middle, and squeezed gently on the micropylar end, resulting in the liberation of the egg, zygote and proembryo from ovules at selected stages. These cells can be used to explore fertilization and embryonic development using molecular biological methods for each cell type and development stage.

中文翻译：

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韭葱（Allium tuberosum Roxb）雄性和雌性配子、合子和原胚的分离

总结雄、雌配子的分离是研究高等植物受精机制的有效方法。渗透压冲击法用于破碎花粉粒马铃薯Roxb 并释放大量收集的花粉内容物，包括生殖细胞。授粉的花柱在 3 小时后被剪掉体内生长，并在培养基中培养6-8小时，在此期间花粉管从花柱的切口端长出。将花粉管转移到含有 6% 甘露醇的溶液中后，管子破裂并释放出成对的精子细胞。胚珠马铃薯在酶溶液中孵育 30 分钟，然后解剖去除珠被。转移到不含酶的解剖溶液后，将每个珠心从中间切开，轻轻挤压珠孔端，在选定的阶段从胚珠中释放出卵、合子和原胚。这些细胞可用于使用分子生物学方法探索每种细胞类型和发育阶段的受精和胚胎发育。