| Penaeid shrimp Marsupenaeus japonicus is an important crustacean species.species in aquaculture of China. However the reseach about the molecular mechanisms of the control of the gonad maturation is poor in the crustacean. Identification of differentially expressed genes in ovary and testis will provide insights into the molecular mechanisms of this important process.Aim to understand gonad development in penaeid shrimp, an ACP (annealing control primer) system which based on mRNA differential display PCR (DDRT-PCR) was adapted to identify genes differentially expressed in ovary and testis at different developing stages. By using 20 pairs of ACP primer, 8 differentially expressed genes were obtained. Two of these genes (thrombospondin, cortical rod protein-2) are ovary-prominent genes. Proliferating cell nuclear antigen (PCNA) and ubiquitin-conjugating enzyme E2r (UBE2r) is expressed in testis with higher level than in ovary. Other differential display sequences identified in this study are unknown unique genes.The full-length cDNA sequence of PCNA and UBE2 r was obtained using 5′rapid amplification of cDNA ends ( 5'-RACE) . The nucleotide sequence obtained was 923 bp in length, including 75 bp of 5'untranslated region (UTR), 783 bp of open reading frame, and 65 bp of 3'UTR (excluding the poly(A)+ tail).Analysis of the deduced protein sequence of penaeid shrimp PCNAshowed that it contains 260 amino acids The nucleotide sequence of UBE2r obtained was 1477 bp in length, including 651 bp of 5'untranslated region (UTR), 726 bp of open reading frame, and 85 bp of 3'UTR (excluding the poly(A)+ tail). Analysis of the deduced protein sequence of penaeid shrimp UBE2r showed that it contains 241 amino acids.The real time PCR system with specific primers designed against different expression genes was used to validity the ACP results and to reveal the expression patterns of these ovary and testis differential expression genes. The Real time PCR results supportes ACP results and further showed that the genes expression level changed significantly in ovary and testis which corresponds with their functions in testis and ovary development. |
