河川沙塘鳢GHR和IGF-2基因克隆及mRNA的时序表达  

Cloning and temporal expression analysis of GHR and IGF-2 in Odontobutis potamophila

作　　者：朱文旭 张宏叶 王涛 王丹[1] 张红燕 尹绍武[1] 陈树桥 周国勤 ZHU Wen-xu;ZHANG Hong-ye;WANG Tao;WANG Dan;ZHANG Hong-yan;YIN Shao-wu;CHEN Shu-qiao;ZHOU Guo-qin(Key Laboratory of Biodiversity and Biotechnology of Jiangsu Province, College of Life Sciences, Nanjing Normal University, Nanjing Jiangsu210023, China;Institute of Fisheries Science of Nanjing City, Nanjing Jiangsu210036, China)

机构地区：[1]南京师范大学生命科学学院,江苏省生物多样性与生物技术重点实验室,南京210023 [2]南京市水产科学研究所,南京210036

出　　处：《海洋渔业》2019年第4期421-433,共13页Marine Fisheries

基　　金：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：生长激素/胰岛素样生长因子-轴(growth hormone/insulin-like growth-axis,GH/IGF-轴)是调控鱼类生长的主要内分泌轴线。为探讨生长相关基因GHR、IGF-2对河川沙塘鳢(Odontobutis potamophila)生长发育的调控机制,采用cDNA末端快速扩增(RACE)和实时荧光定量PCR等技术,对河川沙塘鳢的GHR、IGF-2基因进行了克隆和表达模式分析。研究结果显示:河川沙塘鳢GHR基因的cDNA全长序列为2 179 bp,开放阅读框为1 830 bp,共编码609个氨基酸,IGF-2基因的cDNA全长序列为1 586 bp,开放阅读框642 bp,共编码213个氨基酸。采用qRT-PCR方法检测了GHR、IGF-2基因在9个不同组织(脑、肝、心、肌肉、脾、肠、性腺、鳃、肾)和8个胚胎不同发育时期(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、出膜后3 d)的表达情况。结果表明:IGF-2和GHR基因在所检测的9种组织中均有表达,其中以肝脏、肌肉、性腺、脑中的表达量较高,在胚胎发育阶段GHR和IGF-2基因表达呈先上升后下降的趋势,在原肠胚期表达量较高,表明其在胚胎发育过程中发挥重要作用。qRT-PCR进一步比较了雌、雄河川沙塘鳢GHR和IGF-2基因在不同生长发育阶段(90、150、210、270、330 d)脑、肝、肌肉mRNA的表达量,结果表明:各时期雄鱼GHR和IGF-2基因在肝组织中表达量均显著高于雌鱼,肌肉和脑GHR基因mRNA的表达量则无显著差异。而IGF-2基因则仅在雌、雄鱼210 d的脑和150、210 d的肌肉存在显著性差异,推测肝GHR和IGF-2基因mRNA表达的雌雄差异是河川沙塘鳢雌雄生长差异的主要原因之一。Growth hormone/insulin-like growth-axis(GH/IGF-axis) is the major endocrine axis regulating the growth of fish. In order to reveal the mechanism of male and female growth differences in Odontobutis potamophila, we studied the regulation of growth-related genes GHR and IGF-2 on the growth and development. The rapid amplification of cDNA ends(RACE) approaches and quantitative real-time PCR were used to clone the full-length cDNA of GHR,IGF-2 to analyze their expression patterns. The results showed that the full-length cDNA sequence of GHR gene was 2 179 bp in length, which contained 1 830 bp open reading frame, encoding 609 amino acids. The full-length cDNA sequence of IGF-2 gene was 1 586 bp in length, which contained 642 bp open reading frame, encoding 213 amino acids. qRT-PCR was used to detect the expression of IGF-2 and GHR genes in 9 tissues(brain, liver, heart, muscle, spleen, intestine, gill, kidney) and 8 different development stages from embryo to larva. The results showed that the expression of GHR and IGF-2 genes in embryos rised in the first stage, and then decreased. The expression was the highest in the gastrula, indicating that it played an important role in the process of embryo development. The IGF-2 and GHR expressed in all tissues and expressed higher in liver, muscle, gonad and brain. qRT-PCR was further used to compare the mRNA expression of GHR and IGF-2 genes in brain, liver and muscle at different developmental stages(90, 150, 210, 270, 330 d) between the male and female. The results showed that the mRNA expression of GHR and IGF-2 in males was significantly higher than that in females at each time point. The GHR gene mRNA expression showed no significant difference in muscle and brain, while IGF-2 only showed significant differences between the male and female fish on 210 d and the muscle on 150 d and 210 d. We speculate that the GHR and IGF-2 mRNA variant expression in liver can be accounted for the growth trait differences between male and female Odontobutis potamophila.

关 键 词：河川沙塘鳢 GHR IGF-2 基因克隆 MRNA表达量 

分 类 号：S917.4[农业科学—水产科学]