文章摘要
马晓茜,刘至治,李思发,唐文乔,杨金权.团头鲂主要组织相容性复合体Ⅰ类基因全长cDNA的克隆及组织表达分析[J].上海海洋大学学报,2011,20(1):34-43
团头鲂主要组织相容性复合体Ⅰ类基因全长cDNA的克隆及组织表达分析
Full length cDNA cloning and tissue expression of major histocompatibility complex (MHC) class Ⅰfrom blunt snout bream (Megalobrama amblycephala)
  
DOI:
中文关键词: 团头鲂; 主要组织相容性复合体; cDNA; 克隆  组织表达
英文关键词: Megalobrama amblycephala  major histocompatibility complex(MHC)  cDNA  clone  tissue expression
基金项目:国家自然科学基金重点项目(30630051);上海市重点学科建设项目(S30701)
作者单位
马晓茜 上海海洋大学 水产种质资源发掘与利用教育部重点实验室 
刘至治 上海海洋大学 水产种质资源发掘与利用教育部重点实验室 
李思发 上海海洋大学 农业部水产种质资源与利用重点开放实验室 
唐文乔 上海海洋大学 水产种质资源发掘与利用教育部重点实验室 
杨金权 上海海洋大学 水产种质资源发掘与利用教育部重点实验室 
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中文摘要:
      为了解团头鲂MHCⅠ类基因的结构,采用cDNA末端快速扩增(Rapid-amplification of cDNA ends,RACE)技术,首次成功克隆了团头鲂“浦江I号” F0基础群体及选育F8世代的MHCⅠ类基因,共获得3条cDNA全长序列。序列全长为2 040~2 079 bp,含有87~102 bp的5′-UTR,1 035~1 044 bp的编码区(包括信号肽,alpha 1、alpha 2和alpha 3三个结构域,跨膜区和胞质区)及911~946 bp的3′-UTR区,分别编码347、344个氨基酸。F8世代序列的核苷酸/氨基酸的同源性很高,为89.0%/93.0%,而与F0世代的同源性较低(75.3%~77.0 % 和71.1%~71.4%),呈现出明显的分子多态性。分析表明,团头鲂具有经典的MHCⅠ类分子的空间结构,与人类HLA-A2的抗原肽结合区的晶体结构相比,二者的差异主要集中在5个(I~V)区上。在Neighbor-joining(NJ)法构建的分子进化树中,团头鲂与草鱼的亲缘关系最近,与鲑鳟鱼类、爬行类、鸟类、哺乳类及人类的亲缘关系则渐远。RT-PCR组织表达分析表明,团头鲂MHCⅠ类基因在所检测的的8个组织中均表达,在鳃、头肾和血液中有较强的转录本,中等强度表达于肝脏和脾脏,在后肾、肠和肌肉中表达较弱。
英文摘要:
      RACE (Rapid Amplification of cDNA Ends) technique was used to clone “Pujiang No.1” blunt snout bream (Megalobrama amblycephala) MHC class Ⅰgene (Meam-MHC I), and total three full-length cDNA sequences in the F0 and F8 generations were obtained by cloning and sequencing method, respectively. The sequences were 2 040-2 079 bp in length, including an 87-102 bp 5′-UTR, a 1 035-1 044 bp coding region, containing signal region, three (α1, α2, α3) domains, a transmembrane and cytoplasmic region (TM/CY), and 911-946 bp 3′-UTR. The coding region had 344-347 amino acids. Within F8 generation, the nucleotide/amino acid sequences similarities were as high as 89.0%/93.0%. But the values were relatively as low as 75%-77% /71% between generations, indicating obviously sequence polymorphism. The Meam-MHC I exhibited a classical 3D molecular structure, and compared to the PBR region of HLA-A2,the major differences were mainly in the five regions (I-V). It was revealed in the neighbor-joining tree that the blunt snout bream had the closest relationship with grass carp, far from salmon and trout, and still farther from, chicken and human. Semi-quantitative RT-PCR analysis demonstrated that Meam-MHC I transcript was detected at high level in gill, head kidney and blood, moderate in liver and spleen, and low in trunk kidney, gut and muscle.
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