暗纹东方鲀线粒体COⅡ及两侧tRNA基因的克隆和序列分析

Cloning and Sequence Analysis of Mitochondrial COⅡ and its Connected tRNA in Takifugu fasciatus

用细胞色素氧化酶第二亚基基因(COⅡ)特异性引物对暗纹东方鲀(Takifugu fasciatus)的线粒体DNA(mtDNA)进行PCR扩增,克隆并测定了COⅡ及其侧翼tRNA基因的全序列,结果显示,COⅡ基因691 bp和5'端上游的tRNAAsn基因及3'端下游的tRNALys基因序列共890 bp.用DNA分析软件比较暗纹东方鲀与GenBank中9个目11种鱼类的COⅡ序列,显示暗纹东方鲀与这些鱼类的COⅡ基因具有较高的同源性;其中与同属红鳍东方鲀(T.rubripes)的同源性最高为99.0%.暗纹东方鲀COⅡ基因的核苷酸组成中,A+T含量为56%,与其他11种鱼类的A+T含量(55%～62%)相近.鱼类COⅡ序列组成对A+T核苷酸的偏倚程度比较低.根据暗纹东方鲀与其他11种鱼的COⅡ基因序列同源性所建立的分子进化树,与传统的分类地位基本吻合.推定的tRNA二级结构为典型的三叶草型结构.

Mitochondrial DNA （mtDNA） from the liver tissue of Takifugu fasciatus was prepared as template. The mitochondrial cytochrome oxidase Ⅱgene （ CO Ⅱ） and its connected tRNA genes were amplified by PCR with a pair of specific primers which were designed based on the same gene of T. rubripes from the GenBank. A sequence with a total length of 870 bp including 691 bp nucleotides of COⅡ gene and partial sequences at both sides was cloned. The COⅡ gene of T. fasciatus was proved to have good homology with the other 11 species of fishes recorded in the GenBank and it has 99.0% homology with T. rubripes. The percent of A and T （56%） was similar with that of the other 11 species （55 % - 62 % ）. The pattern of codon usage of the COⅡ gene was less biased. The phylogenetic tree based on the homology of mitochondrial COⅡ gene sequences is basically consistent with that obtained from morphological taxonomic results. Assumed secondary structures of tRNA^Asn and tRNA^Lys genes suggest that they have the classical cloverleaf structures with high rates of complementary base at each arm.