The enzyme dehydrodolichyl diphosphate (dedol-PP) synthase is a cis-prenyltransferase that catalyzes the synthesis of dedol-PP, the long-chain polyprenyl diphosphate used as a precursor for the synthesis of dolichyl phosphate. Here we report the cloning and characterization of a cDNA from Arabidopsis thaliana encoding dedol-PP synthase. The identity of the cloned enzyme was confirmed by functional complementation of a yeast mutant strain defective in dedol-PP synthase activity together with the detection of high levels of dedol-PP synthase activity in the transformed yeast mutant. The A. thaliana dedol-PP synthase mRNA was detected at high levels in roots but was hardly detected in flowers, leaves, stems and in A. thaliana suspension-cultured cells.