河川沙塘鳢DMRT1基因的克隆及其表达分析  

Cloning and expression analysis of DMRT1 gene in Odontobutis potamophila

作　　者：张红燕 朱文旭[1] 赵诚[1] 张宏叶 王涛 陈树桥 尹绍武[1] 贾永义[2] 无 ZHANG Hong-yan;ZHU Wen-xu;ZHAO Cheng;ZHANG Hong-ye;WANG Tao;CHEN Shu-qiao;YIN Shao-wu;JIA Yong-yi(College of Life Sciences,College of Ocean Science and Engineering,Nanjing Normal University, Nanjing 210023,China;Key Laboratory of Healthy Freshwater Aquaculture of Ministry of Agriculture, Key Laboratory of Genetic Breeding of Freshwater Fisheries of Zhejiang Province,Zhejiang Institute of Freshwater Fisheries,Huzhou 313001,China)

机构地区：[1]南京师范大学生命科学学院,海洋科学与工程学院,南京210023 [2]浙江省淡水水产研究所,农业部淡水渔业健康养殖重点实验室,浙江省淡水水产遗传育种重点实验室,浙江湖州313001 [3]南京市水产科学研究所

出　　处：《海洋渔业》2018年第5期569-580,共12页Marine Fisheries

基　　金：浙江省淡水水产研究所农业部淡水渔业健康养殖重点实验室开放课题(重大)(ZJK201803)；江苏省高校自然科学研究面上项目(18KJB240001)；中央财政农业技术推广资金项目(TG15-003)；江苏省科技支撑计划(农业)项目(BE2013441)；南京市科技计划项目(201505059)；常州市科技发展计划项目(CE20162005)

摘　　要：以河川沙塘鳢(Odontobutis potamophila)为研究对象,采用c DNA末端快速扩增(RACE)技术对DMRT1的全长基因进行了克隆,并利用生物信息学技术分析其结构和功能;采用实时荧光定量PCR(qRTPCR)技术检测了河川沙塘鳢DMRT1基因在8种组织(鳃、肠、心、肌肉、脑、肝、脾、性腺)、胚胎发育的8个时期(受精卵期、桑椹胚期、原肠胚期、神经胚期、体节期、口裂期、出膜后1 d和出膜后3 d)以及性腺发育的4个时期(Ⅰ~Ⅳ期)中的表达变化。结果表明,河川沙塘鳢DMRT1基因c DNA序列的全长为2025 bp,编码297个氨基酸,其中包括894 bp的开放阅读框(ORF),73 bp的5'非编码区和1058 bp的3'非编码区。与已知物种的氨基酸序列比对后发现,河川沙塘鳢DMRT1的氨基酸序列与军曹鱼(Rachycentron canadum)、欧洲鲈鱼(Dicentrarchus labrax)的同源性最高。DMRT1基因在河川沙塘鳢的精巢组织大量表达,而在卵巢、肌肉、心以及肝4种组织中表达较少,在其它组织中几乎不表达; DMRT1基因在胚胎发育的各个时期都有表达,在原肠期的表达量最高;此外,DMRT1基因表达量在精巢发育的不同时期呈先下降后上升的趋势,在精子成熟期(Ⅳ期)达到最大值,而在卵巢发育的不同时期表达量较少且表达强度差异不明显,因而推测河川沙塘鳢DMRT1基因与精巢的发生和功能的维持有关。研究结果为解析河川沙塘鳢DMRT1基因的功能及其性别决定机制提供了基础资料,也为开展河川沙塘鳢单性育种奠定了基础。DMRT1 was cloned and characterized from Odontobutis Potamophila with the rapid amplification of cDNA ends(RACE).The expression patterns of DMRT1 in 8 different tissues(gill,intestine,heart,muscle,brain,liver,spleen and gonad)and at different development stages of embryo and gonad(I^IV stage)were detected by real-time fluorescence quantitative PCR(qRT-PCR).The results showed that the full length cDNA of DMRT1 was 2 025 bp,including a 73 bp 5′UTR and a 1 058 bp 3′UTR,894 bp open reading frame(ORF)encoding 297 amino acids.Homological protein analysis showed that Odontobutis Potamophila DMRT1 had higher homology with Rachycentron canadum and Dicentrarchus labrax.The DMRT1 was abundantly expressed in the testis.Except for a slight expression in the ovary,muscle,heart and liver,the presences of DMRT1 were negligible in other selected tissues.The DMRT1 was expressed at all stages of embryonic development and it had the highest expression in gastrula.In addition,the signal of DMRT1 decreased with the process of gonad development and sharply increased at stage IV.However,DMRT1 had no significant variation with the process of ovary development.Therefore,it is speculated that the DMRT1 may be involved in the differentiation and maintenance of testis.The overall expression pattern of DMRT1 in O.potamophila from this study facilitates our understanding of the molecular mechanism underlying sex determination and differentiation,provides valuable information for unisexual breeding of O.potamophila.

关 键 词：河川沙塘鳢 DMRT1基因 QRT-PCR 基因克隆 表达分析 

分 类 号：S917.4[农业科学—水产科学]