We have analyzed an anthracycline biosynthesis gene cluster from Streptomyces nogalater. Based on sequence analysis, a contiguous region of 11 kb is deduced to include genes for the early steps in anthracycline biosynthesis, a regulatory gene (snoA) promoting the expression of the biosynthetic genes, and at least one gene whose product might have a role in modification of the glycoside moiety. The three ORFs encoding a minimal polyketide synthase (PKS) are separated from the regulatory gene (snoA) by a comparatively AT-rich region (GC content 60%). Subfragments of the DNA region were transferred to Streptomyces galilaeus mutants blocked in aclacinomycin biosynthesis, and to a regulatory mutant of S. nogalater. The S. galilaeus mutants carrying the S. nogalater minimal PKS genes produced auramycinone glycosides, demonstrating replacement of the starter unit for polyketide biosynthesis. The product of snoA seems to be needed for expression of at least the genes for the minimal PKS.