Molecular cloning and comparative expression patterns of cyp19a1a of gene in different ploidy cyprinid fishes

In order to study the role of the gonad-specific aromatase(P450aromA)encoded by cyp19a1a gene in ovary development of different ploidy cyprinid fishes,molecular cloning and comparative expression patterns of cyp19a1a of gene in different ploidy cyprinid fishes were carried out.By PCR and rapid amplification of cDNA ends(RACE),the full length cDNAs of cyp19a1a in diploid red crucian carp(Carassius auratus red var.),triploid crucian carp and allotetraploid hybrids were obtained.Our data showed that all the cDNAs of cyp19a1a gene in the three different ploidy fishes encode 517 amino acids,which contains transmembrane helix region,I-helix region,Ozol's peptide region,aromatase-specific conserved region and heme-binding region.By RT-PCR using specific primers against the same sequences of coding regions in different ploidy cyprinid fishes,a comparative study of the expression pattern of cyp19a1awas carried out.This result showed that cyp19a1a was preferentially expressed in ovaries of these three fishes,but also was present at much lower levels in testes,brains and spleens.The expression pattern of cyp19a1a in ovaries of different ploidy fishes from the pre-ovulation and the ovulation was also studied by Real-time PCR,and the results showed that the expression of this gene was greatly different among the three different ploidy fishes.In the ovulation and pre-ovulation,the expression of cyp19a1a mRNA in ovary of triploids was the highest among these three fishes.By immunohistochemistry,the CYP19A peptide locations in the ovary of the different ploidy fishes were studied,which indicated that CYP19A was found in follicle cells,outside layer of zona radiate of Ⅳ oocytes,and ooplasm of stage Ⅱ oocytes.These researches showed that there are different expressions of the gonad-specific aromatase during the ovary development in different ploidy cyprinid fishes.In addition,the abnormal expression of cyp19a1a mRNA in triploids might be associated with sterility.