Saccharomyces cerevisiae S288C produced two laminarinases (1,3-beta-glucanases) which were separated by diethylaminoethyl-Sephadex column chromatography; one was an endo-1,3-beta-glucanase, and the other was an exo-1,3-beta-glucanase active not only on laminarin but also on pustulan (1,6-beta-glucan) and on p-nitrophenyl-beta-D-glucoside. A mutant defective in the production of this last enzyme was isolated, and the mutation was named exb1-1. The selection procedure was based on the capacity of exo-1,3-beta-glucanase to hydrolyze synthetic glucosides. The level of endo-1,3-beta-glucanase in cell extracts of the mutant was normal, but the exo-1,3-beta-glucanase could not be detected by column chromatographic analysis of these extracts. The mutant phenotype, recessive in heterozygous diploids, was stable through successive meioses and showed a Mendelian segregation, indicating that the mutation affected a single gene, which was named EXB1. The lack of production of exo-1,3-beta-glucanase persisted through all the phases of growth, but growth itself was not impaired by the enzyme deficiency.