The Candida albicans ERG26 gene encoding the C-3 sterol dehydrogenase (C-4 decarboxylase) was cloned by complementing a Saccharomyces cerevisiae erg26 mutant with a C. albicans genomic library. Sequence analysis showed a 70% identity between the C. albicans and S. cerevisiae ERG26 genes at the amino acid level. Sequential disruption of both copies of the ERG26 gene in the presence of an integrated rescue cassette containing a third copy of the ERG26 gene under the control of the inducible pMAL2 promoter, resulted in cells capable of growing only in the presence of the inducer. The results establish that the ERG26 gene is essential for growth and that inhibitors of the Erg26p may represent a new and highly effective class of antifungal agents.