利用ISSR标记及rDNA-ITS序列分析河北省野生铁线莲的亲缘关系

ISSR and rDNA-ITS Sequence Analysis of the Genetic Relationship of Clematis in Hebei Province

通过ISSR和rDNA-ITS两种分子标记方法对河北省17个类群的野生铁线莲种间亲缘关系进行了鉴定。利用UPGMA法对ISSR扩增的条带结果进行聚类分析,并用PCR法扩增ITS,通过Clustal X,MEGA等软件比较和分析ITS的序列特征,最后对两种分子标记的结果综合分析。结果表明,利用筛选出的ISSR引物扩增出多态性条带104条,多态性百分比为100%,等位基因数(Na)为2.00,有效等位基因数(Ne)为1.3069,Nei’s基因多样性(H)为0.2196,Shannon’s多样性信息指数(I)为0.3675。聚类结果显示,在遗传距离为0.66处,17种野生铁线莲被分为5类;铁线莲的17个铁线莲样本ITS序列长度为543~561 bp,含变异位点87个,信息位点37个,特异性鉴别位点50个。其中,长瓣铁线莲与半钟铁线莲、太行铁线莲与狭裂太行铁线莲遗传距离为0,槭叶铁线莲与其他种的遗传距离较大,亲缘关系较远。

The ISSR and rDNA-ITS sequence analysis were used to identify the genetic relationship of 17 wild species of Clematis L. in Hebei Province. The ISSR amplified result was conducted cluster analysis by UPGMA method. After PCR amplification,the ITS sequence characteristics were compared and analyzed by Clustal X,MEGA and other software. Finally,the results of these two molecular markers method were comprehensively analyzed. The results showed that primer combinations can amplify 104 polymorphism bands with the percentage of polymorphic bands(PPB)of 100.0%. The average observed number of alleles(Na)was 2.00;the number of effective alleles(Ne)was 1.3069;the Nei’s gene diversity(H)was 0.2196;the Shannon’s information index(I)was 0.3675. According to the clustering results,17 species were divided into five groups at a genetic distance of 0.66. Moreover,the length of ITS sequences varied from 543 to 561 bp,including 87 variable sites,37 information sites and 50 singleton specific discrimination sites in the 17 Clematis L. species. The genetic distance between C. macropetala and C. ochotensis,C. kirilowii and C. kirilowii var. chaneti was 0,and the genetic distance between C. acerfolia and others was large and the genetic relationship was far.