Protoplasts from Neurospora crassa.

Weibull (1953) demonstrated that the intact protoplasts are liberated from cells of Bacillus megaterium after digestion of the cell wall by the enzyme lysozyme, and that these structures persist and exhibit many of the activities of the whole cell provided they are maintained in solutions of appropriate osmotic pressure. Since that time free protoplasts have been obtained from many other gram-positive bacteria through the action of lysozyme, whereas osmotically sensitive, protoplast-like bodies, probably possessing the remnants of a cell wall, have been obtained from a number of gram-negative bacteria by a variety of methods. The now extensive literature relating to free bacterial protoplasts and similar structures has been reviewed in detail recently by Weibull (1958). Liberated protoplasts have proved to be of unique value in morphological, biochemical, and genetic studies of a number of the bacteria from which they have been obtained. It seems likely that homologous structures will be of equal value in studies of other groups of microoorganisms. We therefore set out to obtain protoplasts from the filamentous, coenocytic ascomycete, Neurospora crassa. Necas (1956) obtained structures resembling fragmented free protoplasts from Saccharomyces cerevisiae as a result of (a) mechanical pressure exerted upon the cells and (b) partial autolysis of cultures. He reported the growth and ultimate regeneration of a small proportion of these structures. Eddy and Williamson (1957) liberated intact protoplasts from S. cerevisiae and Saccharomyces carlsbergensis by digestion of the cell wall with enzymes from the gut of the snail Helix pomatia. Emerson and Emerson (1958) have obtained protoplast-like structures from strains of N. crassa bearing the mutant gene os (osmotic) by the action of a commercial hemicellulase and