基于CO I和Cytb DNA条形码在鲌属鱼类物种鉴定中的应用
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摘要
为探究线粒体CO I和Cytb基因作为DNA条形码在鲌属鱼类鉴定中的可行性,分别扩增达氏鲌(Culter dabryi)、海南鲌(C. recurviceps)、尖头鲌(C. oxycephalus)、蒙古鲌(C. mongolicus)、拟尖头鲌(C. oxycephaloides)和翘嘴鲌(C. alburnus)六种鲌属鱼类线粒体细胞色素C氧化酶亚基I(Cytochrome C oxidase subunit I,CO I)和细胞色素b(Cytochrome b,Cytb)的基因片段,并对六种鲌属鱼类同源序列的碱基组成、种内与种间遗传距离以及系统进化分别进行分析。结果表明:长度为645 bp的COI基因片段中,A、T、G、C 4种碱基平均含量分别为25.65%、28.18%、18.33%和27.84%;长度为970 bp的Cytb序列片段中,A、T、G、C碱基平均含量分别为27.97%、27.93%、14.53%和29.57%。基于CO I基因序列的种间平均遗传距离为3.54%,种内平均遗传距离为0.17%,种间遗传距离为种内遗传距离的20.82倍;基于Cytb基因序列的种间平均遗传距离为5.92%,种内平均遗传距离为0.18%,种间遗传距离为种内遗传距离的32.89倍。基于COI基因的系统进化关系显示除尖头鲌与拟尖头鲌外,其余4种鲌均形成独立分支,而基于Cytb基因的系统发育关系显示六种鲌属均能独立形成分支,鲌属六种鱼能够进行有效区分,将CO I和Cytb作为DNA条形码进行鲌属鱼类的物种鉴定是可行的,联合使用可以获得更好的鉴定效果。
Culter dabryi,C. recurviceps,C. oxycephalus,C. mongolicus,C.oxycephaloides and C. alburnus were collected and sequenced to evaluate the effectiveness of DNA barcoding based on mitochondrial cytochrome C oxidase subunit Ⅰ(CO I)and cytochrome b(Cytb).The base compositions of homologous sequences,genetic distance in and between species,and phyletic evolution for six different species of Culter were also constructed.All 50 sequences(11 from GenBank)for CO I and Cytb were used to analyzed,respectively. The results showed that the average percentage of A、T、G and C contents of CO I gene sequences were 25.65%,28.18%,18.33% and 27.84%,respectively.While the average percentage of A、T、G and C contents of Cytb gene sequences were 27.97%,27.93%,14.53% and 29.57%,respectively.The average interspecific genetic distance and average intraspecific genetic distance was 3.54% and 0.17% based on the CO I gene.The overall interspecific genetic distance of CO I sequences was 20.82 times higher than intraspecific genetic distance.The interspecific genetic distance and the intraspecific genetic distance was 5.92% and 0.18% based on Cytb sequences respectively.The interspecific genetic distance of Cytb sequences was 32.89 times higher than the intraspecific genetic distance.The NJ phylogenetic tree based on the CO I fragments showed that all individuals of 4 species except C.oxycephalus and C.oxycephaloides could be clustered into their own groups respectively.The results based on the Cytb showed that all six species in this study could be effectively distinguished.DNA barcoding using CO I or Cytb genes were reaffirmed,what is more,combining these genes were more effective for identification of Culter fish species.
Culter dabryi,C. recurviceps,C. oxycephalus,C. mongolicus,C.oxycephaloides and C. alburnus were collected and sequenced to evaluate the effectiveness of DNA barcoding based on mitochondrial cytochrome C oxidase subunit Ⅰ(CO I)and cytochrome b(Cytb).The base compositions of homologous sequences,genetic distance in and between species,and phyletic evolution for six different species of Culter were also constructed.All 50 sequences(11 from GenBank)for CO I and Cytb were used to analyzed,respectively. The results showed that the average percentage of A、T、G and C contents of CO I gene sequences were 25.65%,28.18%,18.33% and 27.84%,respectively.While the average percentage of A、T、G and C contents of Cytb gene sequences were 27.97%,27.93%,14.53% and 29.57%,respectively.The average interspecific genetic distance and average intraspecific genetic distance was 3.54% and 0.17% based on the CO I gene.The overall interspecific genetic distance of CO I sequences was 20.82 times higher than intraspecific genetic distance.The interspecific genetic distance and the intraspecific genetic distance was 5.92% and 0.18% based on Cytb sequences respectively.The interspecific genetic distance of Cytb sequences was 32.89 times higher than the intraspecific genetic distance.The NJ phylogenetic tree based on the CO I fragments showed that all individuals of 4 species except C.oxycephalus and C.oxycephaloides could be clustered into their own groups respectively.The results based on the Cytb showed that all six species in this study could be effectively distinguished.DNA barcoding using CO I or Cytb genes were reaffirmed,what is more,combining these genes were more effective for identification of Culter fish species.
引文
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[20]柳淑芳,陈亮亮,戴芳群,等.基于线粒体COⅠ基因的DNA条形码在石首鱼科(Sciaenidae)鱼类系统分类中的应用[J].海洋与湖沼,2010,41(2):223-232.
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[23] 谢佳燕,李军德,黄钰淞.鲤科鲌属药用鱼类线粒体COⅠ基因的DNA条形码研究[J].中国中药杂志,2013,38(7):943-946.
[24] Smmons M P,Zhang L B,Webb C T,et al.A penalty of using anonymous dominant markers(AFLPs,ISSRs and RAPDs)for phylogenetic inference[J].Mol Phylog Evol,2007,42:528-542.
[25] Schwarzbach A E,Rlcklefs R E.The use of molecular date in mangrove plant research[J].Wetlands Ecol Manag,2001,(9):195-201.
[26]冯晓宇,谢楠,李行先,等.红鲌属3种鱼类线粒体细胞色素b基因片段序列分析[J].西南大学学报(自然科学版),2008,30(10):80-82.
[27] Zhang X G,Zhang H,Xiong B X.Subspecies validity for genus Culter(Teleostei:Cyprinidae)based on morphometrics analysis[J].Environ Biol Fish,2009,86:193-201.
[2]周剑,陈先均.翘嘴红鲌生物学特性及人工繁殖的初步研究[J].四川农业科技,2005,(1):28.
[3]王丹,程庆武,杨镇宇,等.三峡库区鲌属鱼类线粒体COⅠ基因遗传多样性的初步分析[J].水生生物学报,2015,39(5):1054-1057.
[4] Hebert P D N,Cywinska A,Ball S L,et al.Biological identifications through DNA barcodes[J].Proc R Soc B ,2003,270(1512):313- 321.
[5]Hebert P D N,Ratnasingham S,de Waard J R.Barcoding animal life:cytochrome C oxidase sudunit I divergences among closely related species[J].Proc Royal Soc B Biol Sci,2003,270:96-99.
[6]Savolainen V,Cowan R S,Vogler A P,et al.Towards writing the encyclopedia of life:an introduction to DNA barcoding[J].Philos Trans Royal Soc B,2005,360:1805-1811.
[7] 刘璐,孙典荣,李纯厚,等.DNA条形码技术在鲻科鱼类鉴定中的应用[J].中国海洋大学学报,2016,46(11):178-186.
[8]成跃新,周红章,张广学.分子生物学技术在昆虫系统学研究中的应用[J].动物分类学报,2000,25(2):121-133.
[9] Boore J L.Animal mitochondrial genomes[J].Nucleic Acids Res,1999,2(8):1767-1780.
[10]王琦琦,陈丹丹,唐光辉.核桃举肢蛾线粒体COⅠ和Cytb基因片段序列分析[J].西北林学院学报,2016,31(1):145-152.
[11]宫亚运,章群,曹艳,等.基于线粒体COⅠ基因序列的DNA条形码在中国南海绯鲤属鱼类鉴定中的应用[J].海洋渔业,2016,38(2):113-119.
[12]DeSalle R,Egan M G,Siddall M.The unholy trinity:taxonomy,species delimitation and DNA barcoding[J].Philos Trans Royal Soc London B:Biol Sci,2005,360(1462):1905-1916.
[13]梁宏伟.六种鲇形目鱼类线粒体基因组克隆及其系统发育研究[D].陕西杨凌:西北农林科技大学.2012.
[14] Tamura K,Peterson D,Peterson N,et al.MEGA5:molecular evolutionary genetics analysis using maximumlikelihood,evolutionary distance,and maximum parsimony methods[J].Mol Biol Evol,2011,28(10):2731-2739.
[15] Ward R D,Zemlak T S,Innes B H,et al.DNA barcoding Australia′s fish species[J].Philos Trans Royal Soc B:Biol Sci,2005,360(1462):1847-1857.
[16]徐田军,王日昕,王健鑫.舟山海域4种鲷科鱼类线粒体Cytb基因全序列克隆分析[J].南方水产,2010,6(1):30-35.
[17] Kimura M.A simple method for estimating evolutionary rates of base substitutions through comparative studies of nucleotide sequences[J].J Mol Evol,1980,16(2):111-120.
[18] Hebert P D N,Stoeckle M Y,Zemlak T S,et al.Identification of birds through DNA barcodes[J].PLoS Biol,2004,2(10):312.
[19]彭居俐,王绪祯,王丁,等.基于线粒体COⅠ基因序列的DNA条形码在鲤科鲌属鱼类物种鉴定中的应用[J].水生生物学报,2009,33:271-276.
[20]柳淑芳,陈亮亮,戴芳群,等.基于线粒体COⅠ基因的DNA条形码在石首鱼科(Sciaenidae)鱼类系统分类中的应用[J].海洋与湖沼,2010,41(2):223-232.
[21] Holmquist R.Transitions and transversions in evolution to understanding[J].Mol Phylog Evol,1983,19:134-144.
[22] Avies J C,Walker D.Species realities and numbers in sexual vertebrates:perspectives from an asexually transmitted genome[J].Proc Natl Acad Sci,1999,96(3):992-995.
[23] 谢佳燕,李军德,黄钰淞.鲤科鲌属药用鱼类线粒体COⅠ基因的DNA条形码研究[J].中国中药杂志,2013,38(7):943-946.
[24] Smmons M P,Zhang L B,Webb C T,et al.A penalty of using anonymous dominant markers(AFLPs,ISSRs and RAPDs)for phylogenetic inference[J].Mol Phylog Evol,2007,42:528-542.
[25] Schwarzbach A E,Rlcklefs R E.The use of molecular date in mangrove plant research[J].Wetlands Ecol Manag,2001,(9):195-201.
[26]冯晓宇,谢楠,李行先,等.红鲌属3种鱼类线粒体细胞色素b基因片段序列分析[J].西南大学学报(自然科学版),2008,30(10):80-82.
[27] Zhang X G,Zhang H,Xiong B X.Subspecies validity for genus Culter(Teleostei:Cyprinidae)based on morphometrics analysis[J].Environ Biol Fish,2009,86:193-201.