基于进化树、距离和特征的DNA条形码方法研究——以百花山地区草螟科为例

Application of tree-based,distance-based and character-based DNA barcoding methods for the identification of Crambidae species (Insecta: Lepidoptera) in Baihuashan,China

DNA条形码是一种快捷高效的分子鉴定新技术,近年来在动物分类学领域中得到迅速的发展和应用。在条形码的研究中有基于进化树、距离和特征3种常用的分析方法:第1种方法需要构建系统发育树,分析样本在树上的聚类情况;第2种方法依赖于物种种内和种间的序列差异;第3种则是通过一系列的诊断特征位点来鉴定物种。本研究扩增了北京百花山地区14种草螟科昆虫88个样本的线粒体细胞色素C氧化酶亚基Ⅰ(mitochondrial cytochrome c oxidase subunitⅠ,COⅠ)基因片段,分别基于进化树、距离和特征方法进行了分析,以探讨不同DNA条形码方法在草螟科物种鉴定中的可行性。结果表明:在使用邻接法(neighbor-joining,NJ)构建的系统发育树上,14个草螟物种各自聚成一个单系,均被成功区分。基于Kimura双参数模型计算遗传距离得出,种内和种内有一个明显的"barcoding gap",且ABGD软件对样本的划分完全符合形态鉴定结果。在所有的草螟物种中都找到了诊断核苷酸位点,基于特征来鉴定草螟物种的成功率为100%。结果显示,这3种方法对于本研究中所涉及的草螟都具有较好的区分,基于COⅠ基因的DNA条形码可以作为一种有效的工具在草螟科昆虫的物种鉴定中进行应用。

DNA bareoding has recently been proposed as a promising tool for rapid and accurate species identification in a wide range of animal taxa. Three broad methods （ tree-based, distance-based and character-based） are commonly used. The first method requires the recovery of species as discrete clades （monophyly） on a phylogenetic tree. The second method is based on the degree of DNA sequence variation within and between species. The last method characterizes species through a series of diagnostic characters. To explore the feasibility of DNA barcoding based on different methods in the identification of Crambid moths, the COI genes of 88 samples belonging to 14 species of the Crambidae from Baihuashan were analyzed. The results show that samples belonging to the same species cluster in one monophyly and all species were successfully distinguished by the NJ phylogenetic tree. The intraspecific and interspecifie genetic distances were calculated using the Kimura-2-parameter model. An obvious barcoding gap between intraspecific and interspecific divergences for COI genes was found, while the groups automatically defined by ABGD perfectly match the groups defined by morphological characters. Diagnostic nucleotide positions were found in each species and the character-based barcode method successfully identified every species. This study indicates that the species within the Crambidae can be readilly distinguished using these three barcoding approaches based on the COl gene.