文章摘要
任付真,姚韩韩,钱雪骏,林志华,董迎辉.泥蚶胰岛素生长因子1受体基因的cDNA全长克隆及表达分析[J].水产学报,2017,41(1):40~51
泥蚶胰岛素生长因子1受体基因的cDNA全长克隆及表达分析
Cloning and expression analysis of IGF1R gene in Tegillarca granosa
投稿时间:2015-12-13  修订日期:2016-05-26
DOI:10.11964/jfc.20151210197
中文关键词: 泥蚶  Tg-IGF1R基因  克隆  表达分析
英文关键词: Tegillarca granosa  IGF1R gene  cloning  expression analysis
基金项目:国家现代贝类产业技术体系专项(CARS-48);国家水产种质资源平台项目(2015DKA30470);浙江省自然科学基金重点项目(LZ12C19001);浙江省重中之重学科自设课题(ZS2015012)
作者单位E-mail
任付真 浙江万里学院生物与环境学院, 浙江省水产种质资源高效利用技术研究重点实验室, 浙江 宁波 315100
上海海洋大学水产与生命学院, 上海 201306 
 
姚韩韩 浙江万里学院生物与环境学院, 浙江省水产种质资源高效利用技术研究重点实验室, 浙江 宁波 315100  
钱雪骏 上海海洋大学水产与生命学院, 上海 201306  
林志华 浙江万里学院生物与环境学院, 浙江省水产种质资源高效利用技术研究重点实验室, 浙江 宁波 315100  
董迎辉 浙江万里学院生物与环境学院, 浙江省水产种质资源高效利用技术研究重点实验室, 浙江 宁波 315100 dongyinghui118@126.com 
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中文摘要:
      为了研究IGF1R基因在泥蚶生长、发育过程中的调控作用,本研究基于泥蚶转录组文库的EST,利用SMART RACE技术首次克隆获得泥蚶IGF1R基因(Tg-IGF1R)cDNA全长序列,该序列全长为5793 bp,开放阅读框为4638 bp,编码1546个氨基酸。经生物信息学分析发现,该基因5'端非编码区为810 bp,3'端非编码区为345 bp,Tg-IGF1R蛋白分子量为176.01 ku,等电点为6.04,包含4个Ⅲ型纤粘连蛋白结构域,1个半胱氨酸富集区和1个酪氨酸激酶结构域,属于亲水性跨膜蛋白质。氨基酸多重比对显示Tg-IGF1R蛋白序列与人、非洲爪蟾、斑马鱼的相似性分别为45.2%、45.6%和45.4%。利用qRT-PCR技术检测了Tg-IGF1R基因在泥蚶成体不同组织及幼体不同发育时期的表达情况,发现该基因具有广泛的组织表达性,在所检测的6个组织(血液、闭壳肌、斧足、鳃、内脏团和外套膜)中均有不同程度的表达,其中血液中的相对表达量最高,并与其余5个组织间存在显著性差异,推测Tg-IGF1R以信号分子的形式参与多种组织细胞的生命过程。Tg-IGF1R基因在泥蚶个体发育过程中的9个时期均有表达,且高表达主要集中在原肠胚期,其次是担轮幼虫期和稚贝期,说明Tg-IGF1R基因可能在早期发育阶段参与了某些器官的形成。
英文摘要:
      In order to explore the function of IGF1R during the process of the growth and development in Tegillarca granosa, Tg-IGF1R cDNA was cloned by SMART RACE (Rapid Amplification of cDNA Ends) techniques on the basis of cDNA library of T.granosa. The full length cDNA of Tg-IGF1R was 5793 bp, and its complete ORF (Open Reading Frame) was 4638 bp encoding 1546 amino acids. Bioinformatics analysis showed that the sequence of Tg-IGF1R contains a 5'-untranslated region (UTR) of 810 bp, and a 3'-UTR of 345 bp. The predicted molecular mass of the deduced amino acid of Tg-IGF1R was 176.01 ku, and the theoretical isoelectric point was 6.04. Tg-IGF1R protein consists of four fibronectin type Ⅲ(FN3) domains, one furin like cysteine-rich (FU) domain and Tyrosine kinase (TyrKc) domain, belonging to transmembrane and hydrophilic protein. Homologous analysis on the amino acids showed that IGF1R of T. granosa shared more than 40% similarity with the most species of IGF1R, including Homo sapiens, Xenopus laevis, and Danio rerio. qRT-PCR (Real-time quantitative PCR) results indicated that Tg-IGF1R gene was expressed in 6 tissues (blood, adductor muscle, foot, gill, visceral mass and mantle.), and the expression of blood was significantly higher than other tissues, which implied that Tg-IGF1R plays various biological roles. The results of relative expression in 9 development stages revealed that the expression of Tg-IGF1R gene showed the highest expression in the gastrulae stage, and followed by trochophore and juvenile clams stages. Therefore, Tg-IGF1R may be involved in the formation of certain organs in early developmental stages.
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