浙江乐清铁皮石斛rDNA ITS序列的克隆及序列比对

为判断浙江乐清铁皮石斛（Zhejiang Dendrobium officinale）与石斛属（Dendrobium）其它物种的亲缘关系,本文提取了该石斛鲜样的DNA,利用真核生物ITS序列通用引物ITS4/ITS5进行了扩增,并将扩增产物连接转化至大肠杆菌,在对阳性克隆测序。将得到的序列与GenBank上我国石斛属12个组中34个种的rDNAITS进行了比对,并在此基础上构建了系统发育树。序列比对结果表明浙江乐清铁皮石斛的rDNAITS与黄石斛（D.tosaense）一致,其次与铁皮石斛（D.officinale）最相近,序列相似性99%。系统发育树也表明浙江乐清铁皮石斛的rDNAITS序列与黄石斛（D.tosaense）的相似程度高于其与铁皮石斛（D.officinale）的相似程度。本研究将为利用ITS序列鉴别石斛物种种属关系提供参考。

Cloning and Sequence Alignment of rDNA ITS Sequences of Dendrobium officinale Cultivated in Zhejiang Province

To estimate the relationship between Zhejiang Dendrobium officinale（Leqing） and other Dendrobium spp.,in this paper,we extracted the totall DNA from fresh Zhejiang D.officinale samples and carried on the PCR by using eukaryotic common primer ITS4/ITS5.Then the PCR products were cloned and transformed into E.coli and the positive clones were sequenced.We compared the obtaining sequence with the rDNA ITSs sequences of 34 species among 12 groups of Dendrobium in China which was submitted in GenBank.Based on the alignment,we constructed a phylogenetic tree.The alignment results showed that the rDNA ITS sequence of Zhejiang D.officinale was the same as D.tosaense and the following one was D.officinale with the sequence similarity of Zhejiang D.officinale and D.officinale was 99%.The phylogenetic tree also showed that the level of similarity between Zhejiang D.officinale rDNA ITS and D.tosaense was higher than that of D.officinale.The results of this study would provide a reference for identifying the species and genus relationship of Dendrobium by using ITS sequences.