摘要
【目的】掌握达氏鲟微管相关蛋白1A/1B轻链3B基因(MAP1LC3B)的表达模式,为后续研究MAP1LC3B基因功能及揭示达氏鲟的抗病分子机制提供理论依据。【方法】采用RACE技术从达氏鲟组织中克隆MAP1LC3B基因,通过ProtParam、TMpred、Phyre2及Singal 4.1等在线软件进行生物信息学分析,同时借助实时荧光定量PCR检测MAP1LC3B基因在达氏鲟不同组织中的表达情况。【结果】达氏鲟MAP1LC3B基因cDNA序列全长2208 bp,包含378bp的开放阅读框(ORF)、202 bp的5’端非编码区(5’-UTR)和1628 bp的3’端非编码区(3’-UTR),编码125个氨基酸。MAP1LC3B氨基酸序列含有GABARAP泛素结构域、Agt7结合位点、脂化位点和维管束蛋白结合位点。达氏鲟MAP1LC3B蛋白由18种氨基酸组成,其中谷氨酸(Glu)含量最高(占9.6%)、丙氨酸(Ala)含量最低(占1.6%);蛋白分子量为14743.01 Da,理论等电点(pI)为8.92,不稳定系数为65.12,总平均亲水性为-0.484,是一种不稳定的疏水性蛋白,且不存在跨膜结构,也无信号肽序列。MAP1LC3B基因在达氏鲟鳃、脑、皮肤、头肾、前肾、脾脏、中肾、心脏和肠道等组织中均有表达,以在鳃和肠道组织中的相对表达量较高,显著高于在其他组织中的相对表达量(P<0.05)。【结论】达氏鲟MAP1LC3B基因具有较高的保守性,在鳃和肠道黏膜免疫组织中高表达,表明自噬参与达氏鲟黏膜免疫过程,调节机体免疫反应。
【Objective】The purpose of present study was to clone microtubule-associated protein 1-light chain 3B gene(MAP1LC3B)from Dabry’s sturgeon(Acipenser dabryanus)and analyze its tissue expression,and lay the foundation forstudying the function of MAP1LC3B gene and revealing the molecular mechanism of disease resistance of Dabry’s sturgeon.【Method】The MAP1LC3B gene from A. dabryanus tissue was cloned by RACE technique and analyzed by online bioinformatics softwares ProtParam,Tmpred,Phyre2,Singal 4.1. The expression of MAP1LC3B gene in A. dabryanus tissues was analyzed by real-time fluorescent quantitative PCR(RT-PCR)technique.【Result】The full-length of MAP1LC3B gene of A. dabryanus was 2208 bp. Its open reading frame(ORF)region was 378 bp,in addition,the 5’noncoding region(UTR)and 3’UTR were 202 bp and 1628 bp respectively,encoded 125 amino acids. MAP1LC3B amino acid sequence contained GABARAP domain,Agt7 binding sites,lipidation sites and tubulin binding sites. MAP1LC3B protein consisted of 18 amino acids with highest proportion of glutamic acid(Glu)(9.6%)and lowest proportion of alanine(Ala)(1.6%).Its molecular mass(MM)was 14743.01 Da and the the oretical isoelectric point(pI)was 8.92. Its instability coefficient was 65.12,and the total average hydrophilicity was-0.484,which was an unstable hydrophobic protein. In addition,there was no transmembrane structure and signal peptide sequence. The expression of MAP1LC3B gene was detected in the tissues including gill,brain,skin,head kidney,forekidney,spleen,kidney,heart and intestinal tissues,and the expression of MAP1LC3B was high in the gill and intestinal tissues,and was significantly higher than in other tissues(P<0.05).【Conclusion】MAP1LC3B gene is conservative,its high expression is found in gill and intestinal mucosal immune tissues,suggesting that MAP1LC3B gene may be related to immunity,and participate in the mucosal immunity to regulate the immune response of A. dabryanu.
作者
胡伟
许巧情
郭慧芝
李由申
韩盼盼
袁汉文
张书环
陈敦学
HUWei;XU Qiao-qing;GUO Hui-zhi;LI You-shen;HAN Pan-pan;YUAN Han-wen;ZHANG Shu-huan;CHEN Dun-xue(Yangtze River Fisheries Research Institute,Chinese Academy of Fishery Sciences/Key Laboratory of Freshwater Biodiversity Conservation,Ministry of Agriculture and Rural Affairs,Wuhan 430223,China;School of Animal Science,Yangtze University,Jingzhou,Hubei 434020,China;School of Animal Science,Guizhou University,Guiyang 550025,China)
出处
《南方农业学报》
CAS
CSCD
北大核心
2020年第2期445-452,共8页
Journal of Southern Agriculture
基金
国家自然科学基金项目(31460689)
农业农村部淡水生物多样性保护重点实验室开放项目(LFBC0905)
贵州省科技联合基金项目(黔科合LH字〔2016〕7455号)。
