摘要
目的:通过分析鸦胆子及其混伪品的ITS2序列,探究鉴定鸦胆子药材及其混伪品的新方法。方法:采用改良的CTAB法提取鸦胆子及其混伪品的基因组DNA,PCR扩增ITS2片段。对其进行双向测序,应用MEGA6.0软件进行序列分析,计算种内、种间遗传距离,构建邻接树。结果:鸦胆子药材ITS2序列长度为230 bp,种内最大K2P遗传距离为0.018,小于其与混伪品的种间平均K2P遗传距离0.493。NJ树结果表明鸦胆子序列和柔毛鸦胆子序列聚为一支,与其他混伪品可明显区分。鸦胆子与柔毛鸦胆子在35位点处有一变异位点,利用此位点能将鸦胆子与柔毛鸦胆子区别开来。结论:ITS2可作为鸦胆子药材及其混伪品鉴定的DNA条形码序列,为保障临床用药安全提供了新的技术手段。
Objective：To explore a new method to identify Brucea javanica（L.）Merr.from its adulterants by the ITS2 regions.Methods：Genomic DNA of B.javanica and its adulterants were extracted by modified CTAB method and then the ITS2 sequences were obtained by PCR amplification and sequenced bidirectionally.All of the 41 ITS2 sequences were aligned through Clustal-W and the genetic distances were computed using MEGA 6.0 in accordance with the Kimura 2-parameter（K2P）model.Results：The sequence lengths of ITS2 regions of B.javanica were 230bp.The intra-specific genetic distances were smaller than the average inter-specific ones in ITS2 regions of B.javanica and its adulterants.The NJ tree showed that B.javanica and its adulterants could be easily differentiated according to their monophyly except B.mollis.The ITS2 sequences of B.javanica and B.mollis had a variation point at thirty-fifth site，so B.javanica and B.mollis were able to be identified by this site.Conclusion：ITS2 can be used to identify B.javanica and its adulterants，which provide a new technique to ensure its clinical safety.
出处
《中国现代中药》
CAS
2015年第10期1004-1007,1013,共5页
Modern Chinese Medicine
基金
国家科技支撑计划(2011BAI07B08)
广西自然科学基金项目(2013GXNSFAA019120)
