太湖新银鱼m tDNA COII和Cytb基因的克隆与序列分析

设计了5对特异性引物,扩增、拼接并测定出太湖新银鱼线粒体tRNAAsp-COII-tRNALys和tRNAGlu-Cytb-tRNAThr两段基因序列片段。基因定位和序列分析发现,太湖新银鱼线粒体COII基因全序列长度为691 bp,序列AT含量为52.80%,编码230个氨基酸;线粒体Cytb基因序列全长为1141 bp,AT含量为48.90%,它编码380个氨基酸。分别位于线粒体COII和Cytb基因两翼的4个tRNA基因(tRNAAsp、tRNALys、tRNAGlu和tRNAThr)同时被测定出来。将太湖新银鱼与有明银鱼、小齿日本银鱼的同源序列进行比对分析,并基于线粒体COII Cytb基因合并数据的核苷酸和氨基酸两种序列形式,以黑斑蛙为外群,对10种鱼类进行分子系统树的构建,结果一致表明:小齿日本银鱼与有明银鱼的亲缘关系近于太湖新银鱼;鲱科与鲑科的亲缘关系近于银鱼科鱼类;此外在本研究硬骨鱼类的4个科中,白鲟科作为原始而古老的类群,是在系统进化的过程中首先分化出来的一支。

Cloning and Sequence Analysis of Mitochondrial COII and Cytb Genes of Neosalanx taihuensis

The complete sequences of mitochondrial COII and Cytb genes and their connected tRNA genes(tRNAAsp,tRNALys,tRNAGlu and tRNAThr) were amplified and determined for N.taihuensis by PCR with five pairs of specific primers.The region tRNAAsp-COII-tRNALys of N.taihuensis was 863 bp in length containing the 691-base COIIgene,tRNAAsp gene(73 bp) and tRNALys genes(75 bp),and the region tRNAGlu-Cytb-tRNAThr including the Cytb gene (1141 bp),the tRNAGlu gene(69 bp) and the tRNAThr gene of 72 nt.Two segments of N.taihuensis including the mitochondrial COIIand Cytb genes have been deposited in GenBank under the accession number EF151003 and EF490211,respectively.The authors aligned the COIIand Cytb genes sequences of N.taihuensis with S.ariakensis and S.microdon(the published sequences in GenBank,NC_006918 and NC_004599,respectively) using Clustal X software program.With comparison compared among three sequences of Salangidae,the divergences and base substitutions were analyzed with the MEGA3.1 software.To further confirm the phylogenetic relationships among 3 species of Salangidae,another 7 mt genomes of fishes(S.salar,U12143;O.mykiss,DQ288271;D.cepedianum,NC_008107;S.melanostictus,NC_002616;P.gladius,AY571339;R.porosa,NC_007173 and H.francisci,AJ310141)were retrieved from GenBank,R.nigromaculata(AB043889)being used as the outgroup species.DNA molecular phylogenies were derived using MEGA3.1,with neighbor-joining analyses,based on two data sets combining COIIwith Cytb genes sequences at the nucleotides and amino acid level.The phylogenetic analyses results showed that two trees of NJ both strongly supported a close relationship between S.ariakensis and S.microdon and favored a Salmonidae Clupeidae clade with high bootstap support values.In addition,our results also suggested that Polyodontidae was the most ancestral lineage among 4 families of Actinopterygii.