草鱼ROCK1基因全长克隆、生物信息学分析及组织差异表达  

Full-Length Cloning,Bioinformatics Analysis and Tissue Differential Expression of Grass Carp ROCK1 Gene

作　　者：潘雯倩 汪军鹏 甘甜[1] 胡静[1] 魏静[1] 土志涵 冷向军[1,2,3,4] 李小勤 Pan Wenqian;Wang Junpeng;Gan Tian;Hu Jing;Wei Jing;Tu Zhihan;Leng Xiangjun;Li Xiaoqin(College of Fisheries and Life Science, Shanghai Ocean University, Shanghai, 201306;Key Laboratory of Freshwater Fishery Germplasm Resources, Ministry of Agriculture, Shanghai Ocean University, Shanghai, 201306;Shanghai Engineering Research Center of Aquaculture, Shanghai, 201306;Shanghai University Knowledge Service Platform, Aquatic Animal Genetic Breeding Center, Shanghai Collaborative Innovation Center, Shanghai, 201306)

机构地区：[1]上海海洋大学水产与生命学院,上海201306 [2]上海海洋大学农业部淡水水产种质资源重点实验室,上海201306 [3]上海市水产养殖工程技术研究中心,上海201306 [4]上海高校知识服务平台、水产动物遗传育种中心上海市协同创新中心,上海201306

出　　处：《基因组学与应用生物学》2018年第5期1868-1878,共11页Genomics and Applied Biology

基　　金：上海市科学技术委员会项目(No.13ZR1419600)资助

摘　　要：ROCK1是ROCK（即Rho激酶）家族的成员之一,ROCK家族是小G蛋白Rho的下游效应因子,被Rho蛋白激活后ROCK1作用于下游效应分子,对其介导的信号通路进行调节,参与细胞增殖、迁移、形态改变等。ROCK1与组织纤维化、癌症、心血管等疾病的发生密切相关。本实验采用c DNA末端快速扩增法克隆草鱼ROCK1基因全长序列,预测并分析其编码的蛋白,采用实时荧光定量PCR法检测草鱼ROCK1基因的组织差异表达。实验首次克隆获得了草鱼ROCK1基因全序列,长4 604 bp,编码1 361个氨基酸;草鱼ROCK1氨基酸序列与金线鲃、斑马鱼、鲤鱼ROCK1的氨基酸序列同源性分别达95%、94%、94%,同源性较高;草鱼ROCK1蛋白分子量158 035.73,理论等电点5.69,为亲水性非分泌蛋白,无信号肽;其二级结构包括α-螺旋（α-helix）、β-重叠（β-strand）和环（loop）。组织差异表达显示草鱼ROCK1基因在各组织中均有表达,其中在脑和血液中表达量最高（p〈0.05）,其次为鳃、心脏、肾脏、脾脏和前肠组织,肌肉和肝脏组织表达量较低,皮肤组织表达量最低,所得结果将为后续研究草鱼组织纤维化奠定基础。ROCK1 is one member of the ROCK（Rho-associated coiled-coil-containing protein kinase） family.ROCK family is the downstream effector of small GTP-binding protein Rho.ROCK1 activated by Rho protein acts on the downstream effector molecules,regulates its mediated signal pathway and participates in cell proliferation,migration,and morphological change,etc.ROCK1 is closely related to tissue fibrosis,cancer and cardiovascular diseases.In this study,the full-length sequence of grass carp ROCK1 gene was cloned by rapid amplification of c DNA end（RACE） method,and its encoding protein sequence was predicted and analyzed.The tissue differential expression of grass carp ROCK1 gene was detected by real-time FQ-PCR.In this study,the complete sequence of grass carp ROCK1 gene was cloned for the first time,with a length of 4 604 bp,encoding 1 361 amino acids.The amino acid sequence of grass carp ROCK1 showed a high homology of 95%,94% and 94% with golden-line barbel（Sinocyclocheilus graham）,zebrafish（Danio rerio） and common carp（Cyprinus carpio） respectively.The molecular weight of grass carp ROCK1 protein was 158 035.73,and the theoretical isoelectric point was 5.69.It was a hydrophilic and non-secretory protein with no signal peptide,and its secondary structure included α-helixs,β-strand and loop.The grass carp ROCK1 gene was expressed in all tissues,the maximum expression was found in the brain and blood（p〈0.05）,followed by the gills,heart,kidneys,spleen,and foregut.The expression in the muscle and liver was lower and the lowest expression was in the skin.The results might lay a foundation for further research on tissue fibrosis of grass carp.

关 键 词：ROCK1 草鱼 RACE 基因克隆 组织差异表达 

分 类 号：S917.4[农业科学—水产科学]