基于ITS2序列SNP位点鉴定白及药材及其混伪品  

Identification of Bletillae Rhizoma and its adulterants by SNPs in ITS2

作　　者：赵丹[1] 周涛[1] 江维克[1] 肖承鸿[1] 康传志 

机构地区：[1]贵阳中医学院,贵州贵阳550002

出　　处：《中国中药杂志》2015年第18期3573-3578,共6页China Journal of Chinese Materia Medica

基　　金：贵州省中药现代化科技产业研究开发专项(黔科合ZY字[2013]3001号);贵州省教育厅普通高等学校特色重点实验室建设项目(黔教合KY字[2013]108号);贵州省研究生工作站建设项目(黔教研合JYSZ字[2014]016)

摘　　要：为建立白及药材的分子标记鉴定方法,该文提取白及及其混伪品药材的基因组DNA,利用PCR技术扩增r DNA ITS2片段,片段经双向测序、排序、比对,构建邻接树,查找SNP位点。结果显示,ITS2序列不仅能够有效地区分白及与其混伪品,而且存在可用作白及、黄花白及与其混伪药材鉴别的SNP位点。针对标记白及药材的SNP位点设计引物,通过筛选引物和优化特异性扩增条件后,获取引物BJ59-412F,BJ59-412R,HHBJ-225R在同一扩增条件下,可有效扩增白及、黄花白及,其产物长度分别约为350,520 bp,而混伪品则无PCR条带产生。该文所述引物和建立的反应条件可成功将白及、黄花白及与其混伪品药材进行同步鉴定,操作快速、简捷,结果可靠,可作为白及药材的快速分子鉴定方法。To establish a molecular identification method for Bletillae Rhizoma,this paper extracted genome DNA from Bletillae Rhizoma and its adulterants. The sequences of r DNA ITS2 were sequenced after amplifying. Then multiple alignments of ITS2 were constructed phylogenetic tree with Neighbor Joining by MEGA 5. 1 and found out SNPs loci. The result showed that r DNA ITS2 region could identify Bletillae Rhizoma and its adulterants. There existed the SNPs loci,which could identify Bletilla striata and B. ochracea.Furthermore,we designed specific primers against the SNPs loci of B. striata and B. ochracea,then screened primers and optimized the PCR amplification conditions. Finally,the DNA of B. striata and B. ochracea were specifically amplified by BJ59-412 F,BJ59-412 R and HHBJ-225 R. The length of amplification products were respectively about 350 bp and 520 bp that were effectively identified of B. striata and B. ochracea. While,the adulterants of Bletillae Rhizoma were no-reaction occurring. To sum up,the amplification conditions of the primers can identify B. striata,B. ochracea and their adulterants successfully at the same time. This method was easy,time-saving,and reliable,which can be used as a rapid method for molecular identification of Bletillae Rhizoma.

关 键 词：白及药材 RDNA ITS2 SNP位点 特异性PCR 

分 类 号：R282.5[医药卫生—中药学]