Isolation and purification of generative cells from fresh pollen of Vicia faba L.

A technique named two-step osmotic shock was developed for isolating generative cells (GCs): pollen grains were incubated in 20% sucrose solution and shocked by adding water. This caused the pollen grains to burst and release their contents including GCs. By subsequent filtration and centrifugation the isolated GCs were purified. The viability of GCs before and after purification was confirmed by fluorescein diacetate (FDA) test. This procedure offers a rapid and effective means to obtain living GCs in quantities.