基于ITS2条形码序列鉴定中药材两头尖及其混伪品

Identification of Anemone raddeana and Its Adulterants by ITS2 Sequence

目的:本研究利用ITS2序列对中药材两头尖及其混伪品进行DNA条形码鉴定,以确保该药材的质量及临床疗效。方法:采用试剂盒法对36份样品提取DNA,通过聚合酶链式反应(PCR)扩增其ITS2序列并双向测序,所得序列经过CodonCode Aligner拼接后,用软件MEGA5.1进行数据分析,包括变异位点分析、种内种间距离计算及系统发育树构建。结果:两头尖药材的ITS2序列长度为216 bp,种内最大K2P距离为0.014,种间的最小K2P距离为0.021;NJ树结果显示两头尖与其混伪品可明显区分,表现出良好的单系性。结论:ITS2序列作为DNA条形码能稳定、准确鉴别两头尖药材,为其鉴定提供新的技术手段。

In this study, the internal transcribed spacer 2 of nuclear ribosomal DNA （ITS2） sequence was used for i- dentifying Anemone racldeana and its adulterants to ensure the quality of medicines and clinical efficacy. Genomic DNA was extracted from 36 samples using Genomic DNA kit and used as templates for Polymerase Chain Reaction （PCR）. Sequence assembly and consensus sequence generation were performed by CodonCode Aligner. The intraspe- cific and interspecific genetic distances were computed and the neighbor-joining tree was constructed by MEGA 5.1 in accordance with the Kimura 2-Parameter （K-2P） model. Results： The length of ITS2 sequence of Anemone tad-deana was 216 bp. The Maximum intraspecific genetic distance was 0.014, the minimum interspecific genetic dis- tance was 0.021. The NJ tree showed that A. raddeana differ from its adulterants obviously. Conclusion： ITS2 se- quence was able to identify A. raddeana and its adulterants correctly stably and correctly, which provides a new tech- nique to its identification.