牦牛心脏脂肪酸结合蛋白基因的分子克隆和进化分析

Molecular Cloning and Evolution Analysis of the Yak Heart Fatty Acid-binding Protein Gene

对牦牛心脏脂肪酸结合蛋白(H-FABP)基因进行了克隆测序,并与GenBank中9个物种相应基因编码区核苷酸序列进行了比对分析,在此基础上采用邻接法、最大简约法和最小进化法构建了牦牛与其它物种间分子系统进化树。结果表明,牦牛H-FABP基因由4个外显子和3个内含子组成,外显子1、外显子2、外显子3和外显子4大小分别为73、173、102和54 bp,内含子1、内含子2和内含子3大小分别为3 460、1 892和1 495 bp。CDS序列全长为402 bp,前体氨基酸数为133个。不同物种间在该基因核苷酸序列上有较高的保守性。牦牛与普通牛、绵羊、山羊、猪、人、大鼠、小鼠、鸡、斑马鱼各物种在H-FABP基因编码区核苷酸序列上同源性大小分别为99.8%、97.8%、97.0%、92.8%、88.8%、83.3%、83.1%、76.4%、68.7%。通过邻接法、最大简约法和最小进化法用H-FABP基因编码区核苷酸序列构建的物种间分子系统进化树,结果表明,3种方法构建的物种间分子系统进化树基本一致。系统树总体分为两支,斑马鱼为独立的一支,而牦牛与其它物种为另一大分支。牦牛与普通牛、绵羊与山羊先分别聚在一起,然后再聚为一类;后与猪、人依次聚为一类。小鼠和大鼠先聚为一类,再与人和其它物种聚类,然后再与鸡聚为一类。该系统聚类结果与动物学分类一致,表明H-FABP基因适合于构建不同物种间的系统进化树。

The yak heart fatty acid-binding protein （H-FABP） gene was cloned and compared the nucleotide sequence of coding region with the homologous H-FABP genes in other species including cow, sheep, goat, pig, human, rat, mouse, chick and zebrafish retrieved from the GenBank, and finally a phylogenetic tree was constructed by NJ, MP, ME methods using the coding region DNA sequences of the H-FABP genes in all species. The results showed that the yak H-FABP gene had 4 exons and 3 introns. The size of exon 1, exon2, exon3 and exon4 was 73,173,102 and 54 bp respectively; while the size of intron 1, intron2 and intron3 was 3 460,1 892 and 1 495 bp respectively. The size of CDS sequence of the yak H-FABP gene was 402 bp which encoding 133 amino acids. The nucleotide sequences among different species were quite conservative. The homologies of the coding region of H-FABP genes between the yak and cow, sheep, goat, pig, human, rat, mouse, chick and zebrafish were 99.8%, 97.8%, 97.0%, 92.8%, 88. 8%, 83. 3%, 83. 1%, 76.4%, 68. 7% respectively. The molecular phylogenetic tree among species was constructed according to the nucleotide sequence of the coding region of H-FABP gene. The result indicated that the molecular phylogenetic tree had two branches, branch 1 contained Zebrafish, branch 2 contained yak and other species. Yak and cow, sheep and goat assembled separately, then assembled to a genus, and then assembled to a genus with pig and human, while the rat and mouse assembled to a genus, then assembled to a genus with human, and then chick. This result of phylogenetic clustering was identical to the zoological classification system, indicating that the H-FABP gene was also fit to construct molecular phylogenetic tree among different species.