利用SSR分子标记鉴别柴胡栽培种质的方法学研究

Studies on Identification of Bupleurum Cultivated Germplasm Using SSR Molecular Markers

目的:建立利用简单重复序列(SSR)分子标记鉴别柴胡栽培种质的方法,初步形成可鉴别选育品系的SSR特征谱带数据。方法:从实验室先前设计的柴胡SSR引物序列中选择了50对,以2个选育品系和4份其他柴胡栽培种质为材料,筛选扩增效果好、多态性高的引物对,优化聚合酶链反应(PCR)扩增、电泳等条件,分析获得的SSR谱带,依据遗传距离构建聚类树图。结果:筛选出9对多态性高的引物,明确了检测条件,获得了试验种质的SSR特征谱带数据,在遗传相似系数为0.73处可将所有样品分为4类:黑龙江产红柴胡和川红柴1号品系聚为一类,川北柴1号品系和中柴1号品种聚为一类,四川枫顺和荣县产柴胡种质各独自聚为一类。结论:本研究筛选出的引物对和建立的检测方法可供柴胡种质鉴别参考使用。

This study was aimed to establish the method of identifying bupleurum cultivated germplasm using simple sequence repeat（SSR） molecular markers and to initially establish dataset of characteristic SSR bands to the bred cultivars or strains. From the bupleurum SSR primer pairs which were designed in previous work, 50 primer pairs were selected. Two bred strains and 4 other bupleurum cultivated germplasms were used as test materials. Primers pairs were screened with effective PCR amplification and high polymorphism. Meanwhile, conditions for PCR amplification and electrophoresis were optimized. Then, obtained SSR bands were analyzed and a clustering tree on the basis of genetic distance was constructed. The results showed that 9 SSR primer pairs can be used for identification. The suitable assay conditions were established and characteristic SSR bands were obtained for tested materials. The tested samples can be divided into 4 categories in the genetic similarity coefficient of 0.73. The B. scorzonerifolium cultivated in Heilongjiang and Chuanhongchai No. 1 strains were clustered as one category. Chuanbeichai No. 1 strain and Zhongchai No. 1 cultivar clustered as another category. Cultivated germplasms from Sichuan Fengshun and Rongxian clustered as a unique category. It was concluded that the primer pairs and assay method established in the present study can be used as reference in identification of bupleurum cultivars or cultivated germplasms.