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Transformer-2 (Tra-2)  is an upstream regulatory element of the sex regulation mechanism in insects and plays a critical role in sex formation. To understand the role of  tra-2  in  Hyriopsis cumingii , the full-length  Hctra-2  (1867 bp) was obtained from the gonads, and sequence alignment with other species showed that HCTRA-2 protein had a highly conserved RRM domain. Phylogenetic analysis showed that the HCTRA-2 protein was a close relative to of the mollusks TRA-2 protein. The qRT-PCR of tissue-specific expression pattern showed that the  Hctra-2  was abundant in gonads, and the expression in testes was higher than that in ovaries ( p  < 0.01). It suggests that  Hctra-2  may play a potential regulatory role in gonadal development of  H. cumingii . In the early gonadal development, the  Hctra-2  expression was the highest on the third day after fertilization and increased slightly from 4 months to 5 months, which may be related to the embryonic sex determination and early gonadal development.  In situ  hybridization showed that  Hctra-2  mRNA signals were present in both male and female gonads. After silencing  Hctra-2  by RNAi, the expression levels of  Hcfem-1b  and  Hcdmrt  were changed. It is speculated that there may be a certain relationship between them, which plays an important role in the sex regulation of  H. cumingii . Our research will help to deepen our understanding of the shellfish sex determination mechanisms.

Identification and Functional Analysis of the Sex-Determiner Transformer-2 Homologue in the Freshwater Pearl Mussel, Hyriopsis cumingii