虾夷扇贝Dmrt1基因的分子鉴定及表达模式分析  

Molecular identification and expression pattern analysis of Dmrt1 in the scallop,Patinopecten yessoensis

作　　者：赵丹 周丽青[1,2] 郑言鑫 孙秀俊[2] 吴彪 刘志鸿[2] 吴宙[2,3] 吴磊[2,4] ZHAO Dan;ZHOU Liqing;ZHENG Yanxin;SUN Xiujun;WU Biao;LIU Zhihong;WU Zhou;WU Lei(National Demonstration Center for Experimental Fisheries Science Education,Shanghai Ocean University,Shanghai201306,China;Key Laboratory of Sustainable Development of Marine Fisheries,Ministry of Agriculture and Rural Affairs,YellowSea Fisheries Research Institute,Chinese Academy of Fishery Sciences,Qingdao 266071,China;National Engineering Research Center For Marine Aquaculture,Zhejiang Ocean University,Zhoushan 316022,China;College of Marine Science and Fisheries,Jiangsu Ocean University,Lianyungang 222005,China;Changdao Enhancement and Experiment Station,Chinese Academy of Fishery Sciences,265800,China)

机构地区：[1]上海海洋大学水产科学国家级实验教学示范中心,上海201306 [2]中国水产科学研究院黄海水产研究所农业农村部海洋渔业可持续发展重点实验室,山东青岛266071 [3]中国水产科学研究院长岛增殖实验站,浙江舟山316022 [4]浙江海洋大学国家海洋设施养殖工程技术研究中心,江苏连云港222005 [5]江苏海洋大学海洋科学与水产学院,山东烟台265800

出　　处：《中国水产科学》2021年第5期541-549,共9页Journal of Fishery Sciences of China

基　　金：国家自然科学基金项目(31672637);浙江重中之重开放课题(KF2018008);国家重点研发计划项目(2018YFD0900800)。

摘　　要：Dmrt1是Dmrt双性和mab-3相关转录因子基因家族的重要成员之一,在生殖细胞分化和性别调控中发挥着重要作用。为探究其在虾夷扇贝(Patinopecten yessoensis)性别分化中的表达调控模式,本研究利用生物信息学的方法分析了虾夷扇贝Dmrt1的序列特征;采用半定量PCR(RT-PCR)检测了Dmrt1在不同组织中的表达量差异;应用实时荧光定量PCR(qRT-PCR)和原位杂交技术ISH(in situ hybridization)揭示了Dmrt1在性腺发育4个时期(增殖期、生长期、成熟期、排放期)中的时空表达变化。结果显示,虾夷扇贝Dmrt1序列包含Dmrt基因家族共有的DM结构域;与已知物种同源序列比对后发现,虾夷扇贝Dmrt1序列与欧洲大扇贝(Pecten maximus)同源性最高;原位杂交检测到的阳性信号主要定位在生殖细胞的细胞质中;qRT-PCR定量结果发现,Dmrt1在精巢生长期和成熟期的表达水平显著高于卵巢,在卵巢各发育时期中的表达量无明显变化且维持低水平状态。此外,在外套膜、鳃、肾、和闭壳肌中检测到少量表达的Dmrt1转录本,而在肝胰腺中未检测到。研究结果表明,Dmrt1在虾夷扇贝生殖细胞分化过程中的表达特征与在其他动物性腺发育中的表达特征基本一致,推测其是虾夷扇贝性别分化调控中的关键基因。Dmrt1 is one of the most important members of the Dmrt family and plays an important role in germ cell differentiation and sex regulation.To explore its regulatory expression mode in the sex differentiation of the scallop Patinopecten yessoensis,we analyzed the sequence characteristics of P.yessoensis Dmrt1 using bioinformatics methods.The expression of Dmrt1 in different tissues was detected using semi-quantitative reverse-transcription PCR(RT-PCR).Quantitative fluorescence RT-PCR(qRT-PCR)and in situ hybridization were used to reveal temporal and spatial expression changes of Dmrt1 in the four stages of gonadal development(proliferation stage,growing stage,mature stage,emission stage).The results showed the following:the sequence of P.yessoensis Dmrt1 contains the DM domain shared by Dmrt family;P.yessoensis Dmrt1 sequence had high homology with Pecten maximus,compared with multiple sequences of other known species;the positive signal detected by in situ hybridization,was mainly located in the cytoplasm of germ cells;and qRT-PCR showed that the expression level of Dmrt1 was significantly higher in the testis than in the ovaries during the growing and mature stages.However,there was no significant change in the ovarian development cycle and the expression level of Dmrt1 remained low.In addition,Dmrt1 transcripts were weakly expressed in the mantle,gill,kidney,and adductor muscle but not in the hepatopancreas.The results of the study showed that the expression characteristics of Dmrt1 in gonadal and sex differentiation of P.yessoensis are generally similar to those of other animals.We speculated that Dmrt1 is a key gene for the regulation of sex differentiation in P.yessoensis.

关 键 词：虾夷扇贝 DMRT1 荧光定量PCR 原位杂交 性别分化 

分 类 号：S917[农业科学—水产科学]