ABSTRACT
The production of mutant strains of an organism is particularly useful when studying development, as mutants that are blocked at a certain stage of development may provide data on the developmental process itself. To determine the number of genes whose products are involved in a process it is necessary to carry out complementation analysis, a procedure which normally requires diploid or heterokaryotic tissue. Although protoplasts have been isolated and regenerated from a number of different species of mosses, they have only been used extensively for the study of development in Physcomitrella patens. Protoplasts of P. patens, isolated by the treatment of protonemal tissue with cellulolytic enzymes, regenerate rapidly with high frequency provided they are incubated in osmotically buffered medium and are supported within a matrix such as agar. Regeneration frequencies of 50% are obtained routinely, and frequencies higher than 90% are common.
