The fact that protoplasts of green algae and molds are able to build new cell walls was recognized at the end of the last century (for review see Nečas 1 ). At that time protoplasts were obtained by plasmolysis and mechanical dissection. These techniques could hardly be applied to yeasts. The successful large-scale preparation of yeast protoplasts was achieved first by controlled mechanical disruption of cells, 2 later by autolysis of cell walls, 3 and lately by enzymatic digestion of cell walls (see Volume II, Chapter 5). Regeneration of cell walls in yeast protoplasts, and reproductive ability of walled protoplasts, were first reported by Necas in 1955 for Saccharomyces cerevisiae, 3 , 4 In these experiments protoplasts were cultivated on the surface of solid nutrient media. That method yielded a very low frequency of regeneration (only about 0.1%). Later the rate of regeneration was increased considerably by embedding protoplasts in gelled media. 5