Institute of Medicinal Plant Development (IMPLAD), Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100193, China.
Chin J Nat Med. 2013 Mar;11(2):121-7. doi: 10.1016/S1875-5364(13)60038-2.
The DNA barcoding method was used to accurately and rapidly identify Corni Fructus and its adulterants.
Genomic DNA extracted from Corni Fructus and its adulterants were used as templates. The ITS (internal trascribed spacer) regions were amplified using polymerase chain reaction. Sequence assembly was performed using CodonCode Aligner V 3.5.4. Genetic distances were computed using MEGA V 5.0. Species identification was conducted using neighbor-joining (NJ) trees.
The ITS sequence length of Corni Fructus was 659 bp. The average intra-specific genetic distance of Corni Fructus was 0.005, markedly lower than the inter-specific genetic distance between Corni Fructus and its adulterants (0.357). The ITS2 sequence length of Corni Fructus was 250 bp. No variation was found among the different samples. The interspecific genetic distance of ITS2 between Corni Fructus and its adulterants was 0.571. NJ trees and BLAST results indicated that Corni Fructus and its adulterants can be easily differentiated with monophyly.
ITS/ITS2 regions can accurately and efficiently distinguish Corni Fructus and its adulterants. In addition, the results not only established the foundation for the clinical safety in the utilization of Corni Fructus, but also provided reference for molecular identification of other Chinese herbal medicine and Chinese herbal pieces.
本研究采用 DNA 条码技术，对山茱萸及其混伪品进行快速准确鉴定。
以山茱萸及其混伪品的基因组 DNA 为模板，应用聚合酶链式反应扩增 ITS（内转录间隔区）序列，用 CodonCode Aligner V3.5.4 软件进行序列拼接，用 MEGA V5.0 软件计算遗传距离，用邻接法（NJ）构建系统进化树。
山茱萸 ITS 序列长度为 659bp，种内遗传距离平均为 0.005，显著小于山茱萸与其混伪品之间的种间遗传距离（0.357）；ITS2 序列长度为 250bp，不同样本间无差异，ITS2 种间遗传距离为 0.571。NJ 树和 BLAST 结果均显示山茱萸与其混伪品能够明显区分，具有单系性。
ITS/ITS2 区可准确、高效地区分山茱萸及其混伪品，不仅为山茱萸临床安全用药提供了依据，也为其他中药材及其饮片的分子鉴定提供了参考。
