Abstract:In our study, for a better understanding of the role regarding UBE2r in the development of gonad in crawfish, the full length cDNA of Procambarus clarkii UBE2r gene (Pc-UBE2r) were cloned and characterized. The full-length Pc-UBE2r cDNA sequence obtained was 3671 base pairs (bp) including 554 bp of 5'-UTR and 2 389 bp in 3'-UTR, while containing an open reading frame of 729 bp encoding a protein of 242 amino acids with a predicted molecular weight of 27.6 ku. Bioinformatics analysis indicated that the Pc-UBE2r gene had a conserved domain specified in ubiquitin proteasome family and Cys94 sites. The homology and phylogenetic analysis showed that the deduced amino acid sequence of UBE2r shared high homology in different species and the highest conservation with Marsupenaeus japonicus. Quantitative real-time PCR analysis revealed that Pc-UBE2r was distributed in various tissues, while its expression in the ovary and testis was significantly higher than other tissues (P<0.05). In the development of ovary, the expression of Pc-UBE2r was the lowest in the non-developmental stage, and increased rapidly in the previtellogenic stage. With the maturity of the ovary, the expression of Pc-UBE2r decreased gradually. Meanwhile, during the development of testis, the expression of Pc-UBE2r was proven highest in the spermatocyte generation stage. In our in situ hybridization experimental procedure, the positive signal of Pc-UBE2r was observed uniformly distributed in the oocyte cytoplasm in the non-developmental stage of the ovary. With the development of the ovary, the positive signals of Pc-UBE2r surrounded the oocyte nucleus and follicle cells can be found. In the testis, the positive signals were mainly distributed in the secondary spermatocytes. Besides, Western blotting analysis exhibited that the expression distributions of UBE2r in tissues were significantly different, but remained highl in ovary and testis (P<0.05). Overall, the experimental results demonstrated that Pc-UBE2r may play an important role in the gametogenesis and gonadal development of crawfish.
