| Esox lucius is an economic fish species with Xinjiang characteristics.It is mainly distributed in the Irtysh River Basin in China.Its meat is delicious,rich in a variety of amino acids,widely loved by consumers.This kind of fish grows fast,can adapt to a wide range of water temperature,and has high economic value.Many studies have shown that the female of E.lucius grows faster than the male,and the development of Parthenocarpy culture is helpful to expand the economic benefits of breeding.The genes of Dmrta1,StAR and Wnt4 were cloned by RT-PCR,and their structures were analyzed by bioinformatics software.Quantitative PCR was used to measure the relative expression of the above genes in the eight different tissues of E.lucius.The preliminary study of their roles in gonadal development of E.lucius can provide basic materials for the study of sex determination mechanism and biological function of E.lucius.The results were as follows:(1)The sequence length of Dmrta1 gene was 1408 bp.The open reading frame(ORF)of Dmrta1 gene was 1032 bp,encoding 433 amino acids.Amino acid sequence prediction showed that the Dmrta1 protein had DM and DMA conserved domains.E.lucius was closer to Oncorhynchus mykiss and Oncorhynchus nerka in evolutionary relationship.Dmrta1 gene was expressed in gonad,gill and meat of both sexes,and the expression in testis was significantly higher than that in ovary.At the age of 180 days and 320 days,this gene was highly expressed in the gonads and gills of E.lucius.The above results showed that the expression of Dmrta1 gene in the testis increased during development.(2)The sequence of StAR gene was 1485 bp in length and its open reading frame(ORF)was 864 bp,and encoded 287 amino acids.Amino acid sequence analysis showed that StAR protein had START conserved domain.According to the phylogenetic analysis of StAR protein,E.lucius is more closely related to Salvelinus alpinus and Salmo trutta,but far away from Gallus gallus、Thamnophis sirtalis and Mus musculus.StAR gene was expressed in testis,ovary,head kidney,liver,muscle and brain,and the most prominent level was found in testis,which was significantly higher than that in other tissues including ovary,and showed the expression pattern of gender dimorphism.(3)The sequence of Wnt4 gene was 1549 bp,of which the open reading frame was 1059 bp,encoding352 amino acids.Amino acid sequence analysis showed that Wnt4 protein had WNT1 conserved domain and signal peptide,and there was transmembrane region at 7-29 of amino acid sequence.Phylogenetic analysis showed that the phylogenetic relationship between E.lucius and Myripristis murdjan was the hithermost,and the phylogenetic relationship between E.lucius and Paracentrotus lividus,Azumapecten farreri and Mytilus coruscus was the farthest.Wnt4 gene was expressed in ovary,testis,intestines,meat,gill and other tissues in E.lucius.The expression of Wnt4 gene in ovary was the highest,and the expression of Wnt4 gene in ovary was significantly higher than that in testis.With the development of ovary,the expression of Wnt4 gene decreased.Through the Molecular cloning and expression analysis of sex related genes,it was found that Dmrta1 and StAR genes were related to male sex development,while Wnt4 gene was related to female sex development.Dmrta1 and StAR genes suggest that they play an important role in testis development and maturation,and Dmrta1 gene may play an important role in promoting gill formation in E.lucius.Wnt4 gene is involved in the formation and maintenance of some organs in the early stage of ovarian development.This study can provide a reference for the sex development mechanism of E.lucius,and provide basic materials for the realization of parthenogenetic culture of E.lucius in the future. |
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