In order to gain a better understanding of the origin and evolution of Ogura male-sterile cytoplasm in radish, sequence analysis was conducted for the mitochondrial locus, orfB, using wild and cultivated radishes. The nucleotide sequence of the entire coding and flanking regions of orfB (approximately 1 kb) was determined for six radishes with normal and Ogura cytoplasm and they were classified into three types (types 1-3). The sequence of the 5' flanking region of orfB was further analyzed in 40 additional plants. Irrespective of the category of radish, plants with Ogura male-sterile cytoplasm contained only the type 1 sequence, whereas plants with normal cytoplasm had either type 2 or type 3. The results suggested that the mutational event, which led to the association of orfB with the male-sterile gene orf138, had occurred only once in the history of radishes.