16S rRNA and COI Gene Barcodes Applied for Identification of Ornamental Fishes
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摘要: 应用通用引物测定观赏鱼市场上价值较高的红尾金龙鱼、黄尾龙鱼、青龙鱼、珍珠龙鱼、银龙鱼、神仙鱼、非洲珠宝鱼、接吻鱼、珍珠毛足鲈、蓝钻石孔雀、孔雀鱼、锦鲤等12种观赏鱼的16S rRNA基因和COI基因部分序列。应用DNA序列分析软件，对这些基因进行同源性比较和系统发育分析。通过GenBank中的核酸BLAST和BOLD SYSTEMS数据库对这些基因序列进行比对，结果表明16S rRNA基因和COI基因可以在物种水平作为基因条码对亚洲龙鱼、珍珠龙鱼、银龙鱼和神仙鱼等观赏鱼进行准确的鉴别，而且应用COI基因构建的系统发育树比16S rRNA基因更加接近现有的鱼类分类体系。但对形态十分相似的慈鲷等观赏鱼，以及红尾金龙鱼、黄尾龙鱼、青龙鱼等同一个物种不同品系的观赏鱼依靠16S rRNA基因或COI基因都无法准确鉴定，需要研究进化速度更快的DNA基因条码。Abstract: Partial sequences of 16S rRNA genes and mitochondrial cytochrome oxidase subunit I (COI) genes from 12 highly valuable ornamental fish species were amplified using universal primers. The species included the red-tail golden arowana, the yellow-tail arowana, the green arowana, Scleropages formosus, Scleropages jardinii, Osteoglossum bicirrhosum, Pterophyllum scalare, Hemichromis letourneuxi, Aulonocara stuartgranti, Helostoma temminkii, Poecilia reticulate and Trichopodus leerii. The homology and phylogenetic analyses on the genes were carried out by using DNA sequence analysis software, and compared with the databases at GenBank and Bold Systems. The sequence analysis indicated that the information on 16S rRNA and COI genes might be applicable for distinguishing S. formosus, S. jardinii, O. bicirrhosum and P. reticulatefrom one another. However, the phylogenetic tree constructed with the COI gene would be more closely related to the conventional classification than 16S rRNA. Regardless, the use of 16S rRNA and/or COI gene was not deemed sufficient to differentiate or identify very similar genera or species, such as cichlids or different lines of a same species, nor among the red-tail arowana, the yellow-tail arowana and the green arowana. It was suggested that DNA barcoding with higher evolution rate be studied to overcome the shortcoming of the current method.
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Key words:
- gene barcode /
- ornamental fish /
- species identification /
- 16S rRNA /
- COI
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图 1 基于12种观赏鱼的16S rRNA基因序列的系统发育树
Figure 1. 16S rRNA gene phylogenetic trees of 12 ornamental fish species
图 2 基于12种观赏鱼的COI基因序列的系统发育树
Figure 2. COI gene phylogenetic trees of 12 ornamental fish species
表 1 12种观赏鱼基因GenBank登录号
Table 1. Genbank accessions on 12 ornamental fish species
观赏鱼名称 GenBank登录号 16S rRNA基因 COI基因 非洲珠宝鱼H. letourneuxi KX816035 KX832882 蓝钻石孔雀A. stuartgranti KX816036 KX832884 接吻鱼H. temminkii KX816037 KX832883 孔雀鱼P. reticulata KX816038 KX832885 珍珠毛足鲈T. leerii KX816039 KX832886 神仙鱼P. scalare KX816040 KX832887 锦鲤C.carpio KX866416 KX832888 红尾金龙S. formosus(red-tail golden arowana) KX816041 KX832889 青龙S. formosus (green arowana) KX816042 KX832891 黄尾龙鱼S. formosus(yellow-tail arowana) KX816043 KX832890 珍珠龙S. jardinii KX816044 KX832892 银龙O. bicirrhosum KX816045 KX832893 
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表 2 12种观赏鱼COI基因的遗传距离
Table 2. Genetic distances of COI gene from 12 ornamental fish species
观赏鱼名称 1 2 3 4 5 6 7 8 9 10 11 1.Hemichromis_letourneuxi 2.Helostoma_temminkii 0.396 3.Aulonocara_stuartgranti 0.234 0.353 4.Poecilia_reticulata 0.330 0.311 0.334 5.Trichopodus leerii 0.299 0.270 0.300 0.289 6.Pterophyllum scalare 0.312 0.290 0.281 0.317 0.352 7. Cyprinus_carpio 0.363 0.334 0.349 0.362 0.392 0.335 8.红尾金龙 0.393 0.363 0.332 0.362 0.329 0.344 0.322 9.黄尾龙鱼 0.399 0.366 0.335 0.365 0.335 0.350 0.325 0.003 10.青龙鱼 0.401 0.366 0.339 0.365 0.328 0.347 0.325 0.003 0.003 11.珍珠龙鱼 0.379 0.395 0.401 0.400 0.387 0.400 0.345 0.243 0.248 0.242 12.银龙鱼 0.379 0.369 0.367 0.370 0.357 0.387 0.298 0.276 0.281 0.282 0.290
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表 3 16S rRNA基因和COI基因与数据库相关序列比对结果
Table 3. Sequence alignments on 16S rRNA and COI gene with database
观赏鱼名称 GenBank搜索结果 BoldSystems搜索结果 16S rRNA基因 COI基因 COI基因 非洲珠宝鱼H. letourneuxi H. letourneuxi (99%)；H. bimaculatus(99%)；H. guttatus(99%) H. letourneuxi(100%)；H. saharae (99%)；H. guttatus (99%) H. letourneuxi(100%)；Tilapia guineensis(99.38%)；H. bimaculatus(99.38%)；H. saharae (99.38%) 蓝钻石孔雀A. stuartgranti A. stuartgranti (100%)；Alticorpus geoffreyi(100%)；Maylandia zebra(99%)；Genyochromis mento(99%)；Petrotilapia nigra(99%)；Cynotilapia afra(99%)；Pseudotropheus crabro(99%) A. stuartgranti(100%)；A. geoffreyi(100%)；M. zebra(99%)；G. mento(99%)；P. nigra(99%)；C. afra(99%)；P. crabro(99%) A. stuartgranti(100%)；Metriaclima greyshaki(100%)；Pseudotropheus socolofi(100%)；Pseudotropheus demasoni(100%)；Melano-chromis johannii(100%)；Pseudotropheus elongatus(100%)；Placidochromis milomo(100%)；Maylandia lombardoi(100%)；Maylandia hajomaylandi(100%)；Maylandia callainos(100%)；Labidochromis chisumulae(100%)；P. crabro(100%) 接吻鱼H.temminkii H. temminkii (99%) H. temminkii(99%) H. temminkii(100%) 孔雀鱼P. reticulate P. reticulata(99%) P. reticulate(100%)；P. wingei(100%) P. reticulata(100%) 珍珠毛足鲈T. leerii T. leerii(100%) T. leerii(99%) T. leerii(99.32%) 神仙鱼P. scalare P. scalare(100%) P. scalare(99%) P. scalare(99.85%) 红尾金龙S. formosus S. formosus(99%) S. formosus(100%) S. formosus(100%) 黄尾龙鱼S. formosus S. formosus(100%) S. formosus(99%) S. formosus(100%) 青龙S. formosus S. formosus (99%) S. formosus(99%) S. formosus(100%) 珍珠龙S. jardinii S. jardinii(100%) S. jardinii(99%) S. jardinii(99.85%) 银龙O. bicirrhosum O. bicirrhosum(100%) O. bicirrhosum(100%) O. bicirrhosum(100%)
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