线粒体COI基因条形码在鲿科鱼类物种鉴定中的应用

Species identification of DNA barcoding based on COI gene sequences in Bagridae catfishes

为了探讨线粒体COI基因作为DNA条形码在中国鲿科(Bagridae)鱼类物种鉴定中的有效性,以及系统发育中的适用性,本研究对4属11种鲿科鱼类进行PCR扩增,获得48条线粒体COI基因序列,同时从Gen Bank筛选获得8种鲿科鱼类的12条COI基因序列进行分析。19种鲿科鱼类的COI基因序列特征显示:长度为674 bp的COI序列片段平均碱基组成为24.82%A,30.44%T,27.10%C和17.64%G,碱基组成呈现明显的AT偏倚性(55.26%),具有硬骨鱼类的线粒体COI基因的碱基组成的典型特征。核苷酸位点中有变异位点226个,简约信息位点195个,单一信息位点31个,转换颠换比为3.35。19种鲿科鱼类的种内、种间和属间平均遗传距离分别为0.0041、0.1136和0.1268,种间遗传距离平均为种内遗传距离的27.7倍。在本研究中,鲿科鱼类所有的物种均形成单系,在物种鉴别上与形态学分类结果基本一致,然而鲿科的4个属中只有鱯属形成单系,黄颡鱼属、属和拟鲿属均未形成单系,其进化地位需要进一步研究。线粒体COI基因作为条形码可有效对鲿科鱼类进行物种鉴定,也为鲿科的系统发育提供了参考。

The Bagridae catfishes,belonging to the order Siluriformes and class Osteichthyes,are widely distributed in freshwater areas in Asia,such as China,Japan,Korean peninsula,and Vietnam.In China,Bagridae catfishes include more than 30 species across 4 genera.In recent years,due to environmental deterioration and human disturbance,populations of the native Bagridae had sharply decreased and some of them were difficult to collect.However,the classification of some Bagridae catfishes is ambiguous due to the difference of individuals,genders,or habitat.A species identification technique for various fishes based on DNA barcoding has been rapidly developed and widely employed.For many fishes,DNA barcoding requires additional validation before use.Herein,we have developed and evaluated DNA barcoding based on COI gene sequences for identifying Bagridae species.Altogether,11 Bagridae catfish species were collected and sequenced with the standard DNA barcoding protocols,then combined with 8 available species acquired from the GenBank.A total of 60 sequences from 19 species pertaining to 4 genera were analyzed.DnaSP 5 and MEGA7.0 were employed to calculate the base composition,sequence characterization,and genetic distance,and construct the neighbor-joining phylogenetic tree.The mrbayes software was used to construct the Bayesian tree.The result showed that the mean base contents were:24.82%A,30.44%T,27.10%C,and 17.64%T.AT contents(55.26)were higher and the base composition was biased as others teleost fishes were also commonly observed.The nucleotide sequences included 226 variant sites,195 parsimony informative sites,and 31 singleton sites.The average transition/transversion ratio was 3.35.Genetic distances within species,genus,and families were 0.0041,0.1136,and 0.1268,respectively.The ratio of genetic distances between interspecies and intraspecies was 27.7,which indicated a notable genetic difference of the COI gene sequence.The efficiency of DNA barcoding based on the COI gene was reaffirmed from its successful identification of all 19 Bagridae catfishes in this study.This classification result is consistent with their morphologies.However,only Hemibagrus catfishes can be clustered into a single phylogenetic tree branch.Other Bagridae species were intertwined,and their phylogenetic status needs further research combined with data from more samples.The results revealed that the COI gene was suitable as a DNA barcoding parameter for identifying species;it provided a useful reference for the phylogenetic relationships of Bagridae.