大黄鱼mtDNA ND5和Cytb基因的克隆与序列分析

2004年4月，将采自浙江省象山港海区网箱养殖的大黄鱼样本，提取总DNA，通过设计特异性引物对大黄鱼线粒体DNA（mtDNA）的辅酶5（ND5）和细胞色素b（Cytb）基因进行PCR扩增。扩增产物经琼脂糖电泳检测、纯化后直接测序。得到ND5的序列1839 bp和Cytb基因序列382 bp。应用primer premier5和MEGA3软件包所作的系统发育分析表明：依据大黄鱼ND5序列所作的进化树总体支持传统的分类地位，大黄鱼更接近塘鳢鱼科。而基于Cytb基因所作的分析表明，黑鳃梅童鱼是大黄鱼在石首鱼科中是遗传距离最小的。

Cloning and Sequence Analysis of mtDNA ND5 and Cytb Genes of Large Yellow Croaker Pseudosciaena crocea

NADH dehydrogenase subunit 5 gene(ND5)and cytochrome b gene(Cytb) were cloned in large yellow croaker(Pseudosciaena crocea) cultured in net cages at Xiangshan county,Zhejiang province by PCR with divergent primers in 2004.The products of PCR were sequenced after they were tested and purified with special kits.Both ND5 and Cytb gene had high similarity with other counterpart sequences from BLASTX,and thus these two sequences were useful for phylogenic analysis.The results of sequencing showed that the length of ND5 and Cytb genes were 1839 bp and 382 bp,in which A,T,G and C accounted for 536(29.1%),508(27.6%),236((12.8%)) and 559(30.4%)and 85 bp(22.25%),106 bp(27.75%),64 bp(16.75%)and 127 bp((33.25%)),respectively.Software of Premier 5 and MEGA3 were applied to the phylogenetic analysis,and the phylogentic trees were constructed based on Kimura 2 parameter with other default values.The results indicated that large yellow croaker was close to Ophiocara porocephala based on ND5 phylogeny tree and Collichthys lucidus was the kind of Sciaenidae which had the least genetic distance in Cytb phylogeny tree.The ND5 gene sequence was found to have been sequenced and analyzed,meanwhile the potential of ND5 gene sequence applicable in the phylogeny research in Sciaenidae was studied.The amino acid sequence of ND5 and Cytb were listed in this paper based on previous study and BLASTX similarity search.