Molecular characterization and expression profiles of cdc2 and cyclin B during oogenesis and spermatogenesis in green mud crab (Scylla paramamosain)

摘要

The maturation promoting factor (MPF) is a key regulator of controlling G2/M phase transition in the meiotic maturation of oocyte and spermatocyte in animals, which is a complex of CDC2 (CDK1) and cyclin B. To better understand the molecular mechanism of oocyte and spermatocyte maturation in mud crab (Scylla paramamosain), the full length cDNA of cdc2 (Sp-cdc2) and cyclin B (Sp-cyclin B) were cloned and characterized. The full length cDNA of Sp-cdc2 gene is of 1593. bp encoding a protein of 299 amino acids. Real-time quantitative PCR analysis revealed that the expression level of Sp-cdc2 in the ovary was higher than in other tissues (P < 0.01); and its expression level was not significantly different in different stages of ovary development (P > 0.05), meanwhile there was higher expression in T3 stage than in T1 and T2 stages (P < 0.05). The full length cDNA of Sp-cyclin B is 1492. bp encoding a protein of 391 amino acids. The real-time PCR results showed that its expression level in the ovary was the highest in all examined tissues (P < 0.01), and the gonad expression level in O5 stage was significantly higher than in previous 4 stages and the testis (P < 0.05), and was also significantly higher in T2 stage than in T1 stage (P < 0.05). In situ hybridization analysis showed that the expressions of Sp-cdc2 and Sp-cyclin B transcripts were presented in similar distribution patterns in different developing stages of ovary and testis. The positive signals of Sp-cdc2 and Sp-cyclin B mRNA were detected in the oocytoplasm of oogonia and pre-vitellogenic and primary vitellogenic oocytes, while these two genes had higher expression level in the spermatid and secondary spermatocyte following primary spermatocyte. These results suggested that Sp-cdc2 and Sp-cyclin B may play essential roles in the oogenesis and spermatogenesis of the crab.