暗纹东方鲀性别相关基因dmrt1的分离、鉴定及表达分析  

Isolation,characterization and expression analysis of a sex-related gene dmrt1 in Takifugu obscures

作　　者：高莹莹 黄滨[2] 胡鹏 刘新富[2] 刘滨[2] GAO Yingying;HUANG Bin;HU Peng;LIU Xinfu;LIU Bin(College of Fishers and Life Sciences,Shanghai Ocean University,Shanghai,201306,China;Yellow Sea Fisheries Research Institute,Chinese Academy of Fishery Sciences,Qingdao 266071,China;Neijiang Normal University,Neijiang,Sichuan 641100,China)

机构地区：[1]上海海洋大学水产与生命学院,上海201306 [2]中国水产科学研究院黄海水产研究所,山东青岛266071 [3]内江师范学院,四川内江641100

出　　处：《海洋湖沼通报》2020年第6期99-110,共12页Transactions of Oceanology and Limnology

基　　金：国家重点研发计划(2017YFB0404001);国家海水鱼产业技术体系(CARS-47-G24)资助。

摘　　要：为探究暗纹东方鲀dmrt1基因的序列和结构信息,初步研究dmrt1基因在性腺发育和性别分化过程中的作用,我们利用RACE技术成功分离出暗纹东方鲀dmrt1基因cDNA的全长序列,并对其在成鱼不同组织和幼鱼不同发育期性腺组织中的表达进行了鉴定和分析。结果表明:dmrt1基因cDNA全长1620bp(NCBI登录号:MH218816),编码294个氨基酸,与红鳍东方鲀及星点东方鲀序列相似性最高。dmrt1基因编码的蛋白是一种不稳定的亲水性蛋白,不存在信号肽序列和跨膜区;该蛋白最可能存在于线粒体、细胞质和细胞核;Dmrt1蛋白含有45个磷酸化位点,但不含N(糖基化位点和跨膜保守结构域,二级结构以无规卷曲为主;氨基酸多重序列比对结果显示Dmrt1氨基酸序列的DM结构域在哺乳动物和鱼类中均高度保守,且近C—末端的氨基酸序列也比较保守。荧光定量PCR分析显示dmrt1基因主要在雄鱼中表达,尤其是在精巢中表达量最高,而且dmrt1在幼鱼不同发育期精巢中的表达量呈现出先降低再迅速升高的表达趋势,而相对于精巢而言,卵巢中dmrt仅有微弱表达。研究结果表明,dmrt1可能在雄性性别分化和精巢发育等过程中发挥重要的作用。In order to investigate the sequence and structural information of the dmrt1 gene in Takifugu obscurus, preliminarily study the role of dmrt1 gene in gonad development and sex differentiation, we successfully isolated the full-length cDNA sequence of dmrt1 gene from Takifugu obscurus with RACE technology, subsequently characterized it and analyzed its expression in different tissues of adult fish and gonad tissues of juvenile fish at different developmental stages. The results showed that the full-length of Takifugu obscurus dmrt1 cDNA sequence was 1,620 bp(NCBI accession number: MH218816), encoding 294 amino acids and sharing the highest similarity with that of Takifugu rubripes and Takifugu niphobles. The protein encoded by the dmrt1 was hydrophilic and unstable;it had no signal peptide sequence and transmembrane region. Dmrt1 most likely to exists in the mitochondrion, cytoplasm and nucleus. In this protein, there were 45 phosphorylation sites, but no N-glycosylation site and transmembrane domains. Meanwhile, the secondary structure of this protein was dominated by random coil. The results of multiple amino acid sequence alignment showed that the DM domain of the Dmrt1 amino acid sequence is highly conserved among mammals and fish, and the part of the near C(terminal region in the Dmrt1 was also conservative. Furthermore, the fluorescence quantitative PCR analysis showed that the dmrt1 gene was mainly expressed in male fish, especially in the testis, where it expression level is the highest. The expression of dmrt1 in the testis of juvenile fish at different developmental stages showed a trend of decreasing first and then increasing rapidly, However, dmrt1 was only weakly expressed in ovary as compared with testis. The results showed that dmrt1 may play an important role in male sex differentiation and testicular development.

关 键 词：暗纹东方鲀 DMRT1 cDNA分离 组织表达 发育期 

分 类 号：S917.4[农业科学—水产科学] Q786[生物学—分子生物学]