基于ITS2序列探讨兰属的DNA条形码鉴定和系统发育关系

DNA barcoding identification and phylogenetic relationship in Cymbidium based on ITS2 sequences

兰属中许多类型具有极高的观赏和经济价值,并在兰花产业中占据重要地位。由于兰属植物复杂的演化、遗传历史,一直以来分类鉴定困难,存在诸多分类学争议。近年来,快速发展的DNA条形码技术为兰属植物的分类鉴定提供了新的思路。该研究利用12个兰属本地样品和250条GenBank下载的兰科ITS2序列(其中81条属于兰属),通过BLAST1、遗传变异、建树法等分析评估了ITS2序列用于兰属植物分子鉴定的可行性,并基于建树结果探讨了兰属的系统发育关系。BLAST1结果显示,ITS2序列可以准确鉴定12个本地兰属样品的属、亚属划分,鉴定成功率达到100%;在组水平也具有较好的鉴定能力,鉴定成功率为92%,但在物种水平上的鉴别能力较差,鉴定成功率仅17%。参考库(GenBank下载的250条兰科ITS2序列)遗传变异分析结果显示,兰属ITS2序列具有很高的遗传多样性(变异率为74.9%),在属、亚属水平上能较好地区分,但在组或组下的物种水平,由于组内和组间变异、种内和种间变异存在较大重叠,鉴别能力较差。建树结果显示,ITS2序列可以将兰属中的建兰亚属、兰亚属和大花亚属明显区分开,建兰亚属与兰亚属亲缘较近,两者与大花亚属亲缘较远,同时还发现莲瓣兰与春兰在系统发育树中关系紧密,结果支持Du Puy&Cribb的3亚属划分,并暗示莲瓣兰可能是春兰下的一个变种或品种(支持率为69%)。综上所述,ITS2序列在兰属植物的分子鉴定上具有一定的应用价值,可作为兰属植物DNA条形码鉴定的辅助条形码。

Many types of Cymbidium have high ornamental and economic value and occupy important position in the orchid industry.However,due to the complex evolution and genetic history of Cymbidium,there have been problems in identification and there are many taxonomic disputes.In recent years,the rapid development of DNA barcoding technology has provided new ideas for taxonomy and molecular identification.In this study,12 native samples of Cymbidium and 250 Orchidaceae ITS2 sequences downloaded from GenBank(81 of which belong to the genus Cymbidium)were used to evaluate the feasibility of ITS2 sequence for molecular identification of Cymbidium by BLAST1,genetic variation,and phylogenetic tree-construction methods,and based on the results of phylogenetic tree-construction,explored the phylogenetic relationship of the genus Cymbidium.The results of BLAST1 showed that ITS2 sequence could accurately identify the genus and subgenus of 12 native Cymbidium samples,and the success rate of identification was 100%.It also had good identification ability at the section level,the success rate was 92%.But at the species level,identification ability was poor,and the identification success rate was only 17%.The genetic variation analysis of the reference library(250 ITS2 sequences of Orchidaceae downloaded from GenBank)showed that the Cymbidium ITS2 sequences had high genetic diversity(variation rate was 74.9%),and could be better distinguished at the genus and subgenus levels,but the level of species under the section or section were poor due to the large overlap between intra-and inter-section variation,intra-and inter-species variation.According to the analysis of phylogenetic tree,ITS2 sequence could clearly distinguished subgenus Jensoa,Cymbidium and Cyperorchis.The relationship between the subgenus Jensoa and Cymbidium was relatively close,and they were both far from the subgenus Cyperorchis.At the same time,it was also found that Cymbidium tortisepalum and C.goeringii were close in the phylogenetic tree.This result supported the division of the 3 subgenus of Du Puy&Cribb,and suggested that C.tortisepalum might be a variety or variety from C.goeringii(bootsrap value was 69%).In summary,ITS2 sequences had certain application value in the molecular identification of Cymbidium,and could be used as an auxiliary sequence for the DNA barcoding identification of Cymbidium.