目的 利用DNA条形码进行玉叶金花属植物的分子鉴定。方法 对玉叶金花属20种、89份个体进行叶绿体片段rbcL、matK和trnH-psbA以及核基因片段ITS的PCR扩增和测序,计算种内和种间Kimura 2-parameter(K2-P)遗传距离,利用序列相似性法和邻接法进行单一片段、核心片段、组合片段以及ITS2片段的分析。结果 单一片段分析结果表明玉叶金花属种间遗传距离(0.002～0.012)明显大于种内遗传距离(0.000 3～0.000 7)。trnH-psbA扩增率最低,ITS2片段的分辨率最高,其次是matK和ITS片段,rbcL片段的分辨率最低。片段组合后的分辨率明显提高,基于序列相似性法和邻接法,matK+rbcL+ITS和matK+rbcL+trnH-psbA+ITS的分辨率相当,且比其他片段组的分辨率更高,分别为77%和75%,成功鉴定了15个近缘类群,包括2个药用种类广西玉叶金花和黐花。结论 鉴于trnH-psbA扩增率较低,建议matK+rbcL+ITS作为玉叶金花属物种鉴定的DNA条形码,并作为其药用种类的分子鉴定工具。

Objective This study aims at developing fast and accurate species identification methods for the plants of Mussaenda L. Methods In the present study, DNA barcoding analysis was carried out on 89 individuals representing 20 species of Mussaenda in order to evaluate the performance of the four candidate barcoding loci (matK, rbcL, trnH-psbA, and ITS) and ITS2 region. Based on sequence similarity and Neighbor-joining (NJ) tree reconstruction, we detected inter- and intra-specific genetic distances using Kimura 2-parameter (K2P). Results Inter-specific genetic distance of species in Mussaenda was significantly higher than intra-specific genetic distance. The region of ITS2 showed the highest discrimination power among the independent sequences. Comparably high species discrimination power was also revealed by the matK and ITS data set. The candidate barcode of rbcL displayed the lowest identification rate among the others. However, each individual candidate barcode demonstrated significantly lower discrimination power than the barcode of combined data set. Comparable discrimination power was revealed between the two barcodes of combined sequences matK + rbcL + ITS and matK + rbcL + trnH-psbA + ITS, which showed the values around 77% and 75% based on sequence similarity and NJ tree method. Totally 15 species were identified based on NJ analysis of matK + rbcL + ITS. Conclusion Consequently, the combined sequence of matK + rbcL + ITS provides an effective and fast tool for the identification and authentication of medicinal plant species in the genus Mussaenda L.

国家自然科学基金资助项目(31300174);教育部博士点基金项目(20124404120014