目的 采用目标起始密码子多态性SCoT和内部简单重复序列ISSR 2种分子标记技术对中药材射干、川射干及其混伪品进行鉴定,建立其新的分子鉴别方法。方法 利用优化的ISSR-PCR和SCoT-PCR体系和筛选的引物,对射干、川射干及其混伪品材料进行PCR扩增。利用POPGENE 1.32软件进行多态性分析、利用NTSYS-pc version 2.10e软件构建聚类图。结果 筛选出的10条ISSR引物和SCoT引物,分别扩增出130和143条带,多态性条带百分率分别为96.2%和97.9%。12份材料间的Shannon多样性信息指数(I)分别为0.540 2和0.500 3;Nei′s基因多样性指数(H)分别为0.363 0和0.327 3,表明12份材料间遗传差异较大。基于SCoT标记分析获得的遗传距离高于ISSR标记分析获得的遗传距离。基于SCoT标记聚类分析表明,在相似系数高于0.74时射干、川射干均能独立成支,能较好地与混伪品材料进行区分。结论 SCoT标记能稳定、快速、准确的应用于射干、川射干药材的分子鉴定,ISSR标记仅能用于射干与其混伪品间的分子鉴定。SCoT标记在研究鸢尾属植物种内居群的遗传多样性时比ISSR标记更具优势。

OBJECTIVE To identify Belamcandae Rhizoma, Iridis Tectori Rhizoma and their adulterants by ISSR and SCoT markers.METHODS The genome of Belamcandae Rhizoma, Iridis Tectori Rhizoma and its adulterants were amplified by the optimized ISSR and SCoT PCR conditions and screened primers. Genome polymorphism analysis and the dendrogram construction were calculated by the POPGENE 1.32 and the NTSYS-pc version 2.10 respectively.RESULTS The 130 and 143 bands were amplified by the screened ISSR and SCoT primers respectively. The percentage of polymorphic bands were 96.2% and 97.9% respectively. Shannon diversity index(I) were 0.540 2 and 0.500 3, Nei′s gene diversity index (H) were 0.363 0 and 0.327 3 respectively. The genetic distance calculated based on SCoT marker was higher than that of ISSR marker, which suggested that Elamcandae Rhizoma, Iridis Tectori Rhizoma and their adulterants have great genetic diversity. Cluster analysis based on SCoT genetic similarity indicated that Belamcandae Rhizoma, Iridis Tectori Rhizoma formed independent group with far relationship among their adulterants.CONCLUSION Belamcandae Rhizoma, Iridis Tectori Rhizoma and their adulterants could be distinguished stablely, fastly and clearly by SCoT marker. The ISSR marker could only be used for identification of Belamcandae Rhizoma and its adulterants. The SCoT marker is better suitable for genetic diversity research on the low taxonomic category in genus Iris.