用RAPD技术探讨冬青属苦丁茶的遗传差异、亲缘关系与分类地位

A Study on Genetic Variations,Relationships and Classificationof Several “KUDINGCHA” Species in Ilex(Aquifoliaceae) Based on RAPD Analysis

利用随机扩增多态性DNA (RAPD)技术对 2 2份冬青属苦丁茶不同种质材料的遗传变异进行了研究。 4 0个 10 bp的引物用于多态性检测 ,从中筛选出了 2 7个多态性良好的引物。用该 2 7个引物对上述 2 2份种质材料进行PCR扩增 ,共得到 4 45条DNA谱带 ,其中多态性带4 32条 ,占 97 0 8%。根据扩增结果计算了各份材料之间的遗传距离 ,并用UPGMA构建了聚类树状图。分析结果表明 :2 2份供试材料分成 4类 5组 ,第Ⅰ类为Ilexpentagona (分为A、B两组 :A组为新变种var mashanensisG .M .L .;B组为原变种var pentagona) ,第Ⅱ类为I kudingcha,第Ⅲ类为I latifolia ,第Ⅳ类为I cornuta ;第Ⅱ ,Ⅲ ,Ⅳ三类各有一组材料组成。物种的差异、亲缘关系以及同一物种内不同种质材料在起源地域上的差异均可从系统树中反映出来。RAPD分子标记的结果可作为判断冬青属苦丁茶种质资源材料的起源地域、遗传差异。

The genetic variations,relationships and classification of 4 “KUDINGCHA” species and a few of intermediate-typed materials in Ilex L.of Aquifoliaceae was studied based on RAPD analysis.22 germplasm materials of 4 KUDINGCHA species,i.e.Ilex kudingcha C.J.Tseng,I.latifolia Thunb.,I.pentagona S.K.Chen,Y.X.Feng et C.F.Liang,I.cornuta Lindl.and an intermediate type (it lies between Ilex kudingcha and I.pentagona,in morphology) were analyzed for PCR amplification.27 10-bp random primers with optimal polymorphism were selected from 40 primers tested,and the selected primers were used for the PCR amplification on the 22 germplasm materials mentioned above.445 bands were obtained,of which 432 bands were polymorphic bands,amounting to 97.08% of the total bands amplified.The genetic distances between any two germplasm materials were calculated based on the bands,and the dendrogram was constructed using Nei's genetic similarity values and UPGMA program.The results showed that the 22 germplasm materials can be divided into 4 taxa and 5 groups.Taxon Ⅰ consists of Group A and Group B of pentagona.Group A is a new varietas,var.mashanensis;and Group B is the original varietas,var.pentagona.Taxon Ⅱ,Ⅲ and Ⅳ consist of one Group (one species),respectively.The differences and relationships of species,as well as the differences of origin regions of different germplasm materials of the same species,are clearly reflected in the cluster analysis dendrogram.The results of RAPD analysis can be considered as an important basis for deciding the origin region,genetic variations,and relationship of the various germplasm materials of “KUDINGCHA” in Ilex L.,and for the classification on the species-level or infraspecific level.