三种鳜鱼(Siniperca)生长激素基因的克隆及序列比较

COMPARISON IN CLONING AND SEQUENCE OF GROWTH HORMONE GENE IN THREE SPECIES OF GENUS SINIPERCA

采用长片段PCR及T-A克隆技术,从基因组DNA成功克隆翘嘴鳜、大眼鳜和斑鳜含完整阅读框的生长激素(GrowthHormone,GH)基因DNA全序列,翘嘴鳜序列全长5548bp,大眼鳜序列全长5483bp,斑鳜序列全长5789bp,提交GenBank数据库后获得注册号分别为EF205280、EF205281、EF441623。三种鳜鱼GH基因均由六个外显子、五个内含子组成;均在第二内含子中相同位置发现"AG"微卫星重复序列;均在第三内含子末端发现一处剪接位点序列的相邻重复。三种GH基因在第一、二、三内含子处存在碱基长度差异。序列间差异主要表现为邻近序列的重复、DNA片段的缺失或插入及单核苷酸多态性(SNP)。翘嘴鳜与大眼鳜GH基因编码氨基酸序列完全相同,两者与斑鳜序列在第150位有一个氨基酸残基差异。采用MEGA软件对三种鳜鱼GH基因DNA序列及编码氨基酸序列分别构建UPGMA系统发育树,结果表明,在系统发育中,斑鳜可能最先从共有祖先物种中分化出来。

Growth hormone （GH） genes containing a complete open reading flame were cloned from genomic DNA of Siniperca chuatsi, S. kneri and S. scherzeri using long PCR and T-A cloning techniques. Their lengths were 5548bp, 5483bp, and 5789bp, and the accession numbers of the three GH genes in GenBank are EF205280, EF205281, and EF441623 respectively. All the GH genes were composed of six exons and five introns. An AG repeat microsatellite sequence was found at the same position of the 2nd introns, and a repeated splicing site was found at the end of the 3rd intron in all three genes. The lengths of the 1 st, 2nd and 3rd intron were different among all of these genes. The difference in the sequence was mainly shown in the repetition of nearby fragment, absence or insertion of DNA fragment, and single nucleotide polymorphism （SNP）. The amino acid sequence coded by GH gene of S. chuatsi was the same with that of S. kneri, but it was different from that of S. scherzeri at the 150th loci. Using MEGA software, the UPGMA phylogenetic trees were constructed respectively for DNA sequences of GH genes and the amino acid sequences coded by these genes. The results indicated that S. scherzeri might most probably be first differentiated out from a common ancestor before other two in phyletic evolution.