基于ITS2序列的苍术属植物遗传关系

目的:应用内部转录间隔区2(ITS2)序列鉴定东北产栽培和野生苍术及其近缘种药材,明确其种间遗传关系远近,并对东北产朝鲜苍术的栽培起源进行初步探索。方法:提取不同栽培区包括北苍术、朝鲜苍术在内的五种苍术属40份样本以及朝鲜苍术7份野生种质样本的基因组DNA,通过聚合酶链式反应(PCR)扩增ITS2序列并进行双向测序,所得所有序列用MEGA5. 0软件进行比对分析(clustal W),去除两端5. 8S和28S序列,获得完整的ITS2序列,构建系统聚类树(NJ树)。结果:苍术属5种药材ITS2序列长度均为232 bp。根据NJ树和ITS2二级结构结果,除北苍术和朝鲜苍术未被区分开,其余几种药材均可明显区分,表现出良好的单系性。根据NJ树结果可知,朝鲜苍术的栽培品系和野生种质也能很好的聚在一起。结论:ITS2序列能稳定、准确鉴别苍术属5种药材。朝鲜苍术与北苍术亲缘关系很近,可认为是苍术北方分支中的一种变种,建议将朝鲜苍术并入北苍术。且在辽宁有大规模栽培的朝鲜苍术可能起源于辽宁本地的野生群体,种源可能来自于辽宁岫岩等地的野生种。

Molecular Identification of Atractylodes Plants Based on ITS2 Sequence

Objective: To identify the genetic relationship of cultivated and wild Atractylodes and its closely related species by using the Internal Transcribed Spacer 2(ITS2)barcode,in order to explore the cultivation origin of A. coreana from Northeast China. Method: Genomic DNAs were extracted from 40 samples of Atractylodes and its closely related species from different cultivated habitats,and 7 samples of wild A. coreana. were also extracted. The ITS2 sequences of these samples were amplified, and bidirectional sequencing was conducted by polymerase chain reaction(PCR). Totally 47 ITS2 sequences were aligned by using MEGA 5.0,5.8S and 28S sequences were removed to obtain the complete ITS2 sequence and build neighbor-joining (NJ) tree. Result: The lengths of ITS2 sequences of all samples were 232 bp. The NJ tree and the secondary structures of ITS2 showed that various varieties could be distinguished obviously except A. chinesis and A. coreana,which showed a good monophyly. The NJ tree showed that cultivated and wild A. coreana can also get together very well. Conclusion: As a DNA barcode,ITS2 sequences can be used to stably and accurately distinguish various varieties of Atractylodes. The relationship between A. chinesis and A. coreana is very close. A. coreana can be considered as a variant of A. coreana in North China. It is recommended to incorporate A. coreana into A. chinesis. The large-scale cultivation of A. coreana may originate from local wild population in Liaoning province,and the provenance may come from Xiuyan and other places in Liaoning province.