Sex identification by male-specific growth hormone pseudogene (GH-psi) in Oncorhynchus masou complex and a related hybrid.

It is often difficult to identify sexes of many fish species by conventional cytological method because of the lack of heteromorphic sex chromosomes. Isolation of sex-specific molecular markers is thus important for sexing and for understanding sex chromosome evolution in these species. We have identified genetic sexes by PCR-based male-specificity of a growth hormone pseudogene (GH-psi) in masu and Biwa salmon, two subspecies of the Oncorhynchus masou complex, and their hybrid Honmasu. PCRs with primers designed from sequences of chinook salmon GH genes amplified GH-I and GH-II fragments in both sexes, but a third GH-psi fragment was detected in predominant proportion of males and very few phenotypic females. The consistency of phenotypic sex with genetic sex identified by GH-psi for masu salmon, Biwa salmon and Honmasu was 93.1, 96.7 and 94%, respectively. The remaining individuals showed inconsistency or deviation from sex-specificity: a few phenotypic males lacked the GH-psi, whereas a few phenotypic females possessed the GH-psi. Sequence of the putative GH-psi fragment from such females was identical to that from genetic males, and shared about 95% homology with the corresponding GH-psi fragment from chinook salmon. This result confirmed that these females were really GH-psi-bearing individuals. PCR analyses with primers designed from masu salmon GH-psi gave identical results, indicating that the absence of GH-psi in a few males was not resulted from primer mismatching. These GH-psi-bearing females and GH-psi-absent males were more likely to originate from spontaneous sex reversion than from crossing-over between GH-psi and the sex determination gene/region.