[关键词]
[摘要]
目的 利用ITS2条形码鉴别两面针药材及其混伪品,为两面针药材鉴定提供新方法。方法 采用PCR法扩增ITS2(转录第二间隔区)序列,双向测序后运用CodonCode Aligner、MEGA软件进行数据处理,构建系统聚类树(NJ树)。结果 两面针药材及其混伪品基因组DNA降解明显,但不影响ITS2条形码的PCR扩增和测序,ITS2条形码序列分析表明种内与种间遗传距离具有较大差异,基于ITS2条形码构建NJ树可鉴别两面针药材及其混伪品。结论 ITS2条形码适用于两面针药材及其混伪品的鉴别,为两面针药材快速、准确鉴定提供新方法;为两面针基原研究提供重要的分子鉴定证据;同时为DNA条形码技术应用于其他根、茎类中药材的鉴定提供了示范作用。
[Key word]
[Abstract]
Objective To provide the evidences for molecular identification as well as genetic diversity studies, sequencing and comparison of rDNA ITS sequences from 15 plants in Rubus L. were performed. Methods Full length rDNA ITS sequences were isolated from leaf genomic DNA by PCR method with universal primers, and these sequences was analyzed using bioinformatic softwares. Results The length of ITS1, ITS2, and 5.8 S sequences for the 15 plants in Rubus L. were 255-258, 208-211, and 164 bp, respectively. Total 138 variable sites were found in ITS1 and ITS2 sequences with 41 parsimony information ones, and some transitions, transversions, and deletions were observed. The 5.8 S sequences were more conserved and there were only four variable sites with no parsimony information ones. The genetic distance of the 15 plants in Rubus L. ranged from 0.139 0 to 0.008 1, and the smallest genetic distance was observed between R. irenaeus and R. reflexus. Conclusion The rDNA ITS sequences of the 15 plants in Rubus L. are obtained, which would provide the evidences for molecular identification and genetic diversity studies.
[中图分类号]
[基金项目]
广东省教育部产学研结合项目(2009B090300202)