Two kinds of cDNA (types A and B) encoding human dopamine beta-hydroxylase (DBH) were isolated from a pheochromocytoma cDNA library. Type A (2.7 kb) and B (2.4 kb) encoded the same amino acid sequence and were different only in 3'-untranslated region. Type A contained 3'-extension of 300 bp at the end of type B. Subsequently, we isolated human DBH gene and analyzed genomic DNA by Southern hybridization. Human DBH gene (approximately 23 kb) was composed of 12 exons and existed as a single gene on genome. Exon 12 encoded 3'-terminal region of 1,013 bp of type A, including the 300 bp sequence. These results indicate that alternative use of two polyadenylation sites from a single DBH gene generates different mRNA types. This conclusion was supported by Northern hybridization and S1 nuclease mapping experiments. The ratio of type A and B mRNAs in pheochromocytoma was roughly 1.0 to 0.2. We found possible transcription regulatory elements, TATA, CCAAT, CACCC, GC boxes, near the transcription initiation site of DBH gene. Sequences homologous to glucocorticoid and cyclic AMP response elements were also observed.