新型联合片段:nrDNA ITS+nad 1-intron 2在枫斗类石斛鉴定中的意义

Applylication of new type combined fragments: nrDNA ITS+ nad 1-intron 2 for identification of Dendrobium species of Fengdous

本文从线粒体基因片段入手,利用PCR技术扩增出17种枫斗类石斛39个个体的线粒体基因co I、nad 5部分序列及nad 1-intron 2序列,nr DNA ITS及叶绿体基因序列rbc L、mat K、psb A-trn H,同时筛选出合适的基因片段来研究多片段联合在枫斗类石斛植物鉴定方面的应用。结果表明:在7个基因片段中,nr DNA ITS、nad 1-intron 2和psb A-trn H的序列变异度较高,可用于枫斗类石斛的鉴定。但是,仅利用单一序列无法对细茎石斛及霍山石斛进行鉴别。而nr DNA ITS+nad 1-intron 2新型联合片段构建的UPGMA树与其他联合片段相比,更能有效的对枫斗类石斛基原植物进行鉴别并可区分细茎石斛及霍山石斛。此外,借助nr DNA ITS+nad 1-intron 2新型联合片段构建的UPGMA树对3个待检种进行了成功鉴别,分别为霍山石斛、细茎石斛和铁皮石斛。本研究首次将nr DNA ITS与线粒体基因片段联合用于枫斗类石斛的鉴定,为石斛药材的鉴别提供了可靠的科学依据,同时也将对规范枫斗类石斛市场提供理论指导。

In this study, 17 kinds of Dendrobium species of Fengdous including 39 individuals were collected from 4 provinces. Mitochondrial gene sequences co I, had 5, had 1-intron 2 and chloroplast gene sequences rbcL, matK amd psbA-trnH were amplified from these materials, as well as nrDNA ITS. Furthermore, suitable sequences for identification of Dendrobium species of Fengdous were screened by K-2-P and P-distance. The results showed that during the mentioned 7 sequences, nrDNA ITS, had 1-intron 2 and psbA-trnH which had a high degree of variability could be used to identify Dendrobiurn species of Fengdous. However, single fragment could not be used to distinguish D. moniliforme and D. huoshanense. Moreover, compared to other combined fragments, new type combined fragments nrDNA ITS＋had 1-intron 2 was more effective in identifying the original plants of Dendrobium species and could be used to identify D. huoshanense and D. moniliforme. Besides, according to the UPGMA tree constructed with nrDNA ITS＋had 1-intron 2, 3 inspected Dendrobium plants were identified as D. huoshanense, D. moniliforme and D. officinale, respectively. This study identified Dendrobium species of Fengdous by combined fragments nrDNA ITS＋had 1-intron 2 for the first time, which provided a more effective basis for identification of Dendrobium species. And this study will be helpful for regulating the market of Fengdous.