Two types of cDNA (GTH alpha 1 and -alpha 2) encoding the alpha subunits of masu salmon (Oncorhynchus masou) gonadotrophin were cloned by the reverse transcription-polymerase chain reaction for pituitary mRNAs. The nucleotide sequences showed that the GTH alpha 1 cDNA was 380 bp long, encoding 119 amino acids, and that GTH alpha 2 cDNA was 365 bp long, encoding 114 amino acids. The masu salmon alpha subunit types had a few differences between the sequences, with homologies of 80% (nucleotide sequence) and 72% (amino acid sequence). The structural difference between the alpha 1 and alpha 2 subunits was predicted using hydropathic analysis. The evolutionary interval between masu and chum salmon was estimated to be 4.0 and 2.3 million years by comparing their GTH alpha 1 and -alpha 2 subunits respectively. These time values are roughly consistent with the evolutionary time interval (3.0 million years) estimated from fossil records and an isozyme study. Specific synthetic oligonucleotide probes were constructed and used for genomic Southern blot analyses. The restriction fragment sizes of the GTH alpha 1 and -alpha 2 genes were similar, and when their patterns were compared with those from four other teleosts, each species showed a different pattern from the others, but no difference between their respective alpha 1 and alpha 2 genes. Therefore, the structural features of the GTH alpha 1 and -alpha 2 genes may have diverged in a similar manner in these five teleosts.