The SCS3 gene of Saccharomyces cerevisiae was cloned by functional complementation, using a conditional mutant exhibiting myo-inositol auxotrophy in the presence of choline, and sequenced. The sequence contained an open reading frame capable of encoding 380 amino acids with a calculated molecular weight of 42,734. Disruption of the SCS3 locus caused myo-inositol auxotrophy. The gene appeared to be involved in the synthesis of inositol phospholipids from inositol but not in the control of inositol synthesis.