纤花香茶菜EMS突变体库构建及ISSR与SSR分析

Construction of EMS Mutant Library of Isodon lophanthoides var.graciliflorus and Analysis of ISSR and SSR

为筛选甲基磺酸乙酯(EMS)诱变处理纤花香茶菜种子的最佳处理条件,利用表型鉴定及ISSR和SSR双分子标记技术对典型表型变异植株进行分析,构建纤花香茶菜突变体库,为纤花香茶菜种质创新利用研究提供材料。结果表明,1.2%EMS处理7 h为纤花香茶菜种子最佳诱变处理。利用EMS诱变处理3000粒纤花香茶菜种子,对得到的1262株成株期M 1代材料进行表型鉴定,共筛选出489株表型变异植株,变异频率为38.7%,表型变异性状包括高杆、矮杆、粗茎、细茎、分枝、多分枝、叶色深等。M 2代经表型鉴定初步筛选出4株性状优良的单株。ISSR分析结果表明,9条引物共扩增出72个等位基因位点,平均每条引物扩增出8个位点,多态性位点有54个,多态性比率为75.00%。22个表型变异植株和对照组的遗传相似系数变化范围为0.6389~0.8472;SSR分析结果表明,13对SSR引物共扩增得到86个等位基因位点,平均每条引物扩增出7.81个位点,多态性位点有54个,多态性比率为62.79%。22个表型变异植株和对照组的遗传相似系数变化范围为0.5905~0.8286。基于ISSR及SSR的聚类分析结果都表明,对照组基本上能与表型变异植株区分开,说明大部分诱变植株与对照组在分子水平上具有一定的遗传差异。

This study aimed to identify the optimal treatment conditions for ethyl methanesulfonate(EMS)mutagenesis of Isodon lophanthoides var.graciliflorus seeds and analyze the phenotypic variations of selected mutant plants using ISSR and SSR molecular markers.A mutant library of I.lophanthoides var.graciliflorus was constructed to provide materials for the study of germplasm innovation and utilization in I.lophanthoides var.graciliflorus.1.2%EMS treatment for 7 hours is the optimal mutagenic treatment for the seeds of I.lophanthoides var.graciliflorus.Using EMS mutagenesis treatment,3000 seeds of I.lophanthoides var.graciliflorus were treated.Phenotypic identification was performed on 1262 mature M 1 generation plants obtained.A total of 489 phenotypic variation plants were selected,with a mutation frequency of 38.7%.The phenotypic variations included high straw,dwarf,thick stem,thin stem,branching,multi-branching,and dark leaf color.From the M 2 generation,four individual plants with excellent traits were preliminarily selected based on phenotypic identification.The results of ISSR analysis showed that 9 primers amplified a total of 72 loci,with an average of 8 loci per primer.Among them,54 loci were polymorphic,accounting for a polymorphism rate of 75.00%.The genetic similarity coefficient between the 22 phenotypic variant plants and the control group ranged from 0.6389 to 0.8472.The results of SSR analysis showed that 13 pairs of SSR primers amplified a total of 86 loci,with an average of 7.81 loci per primer.Among them,54 loci were polymorphic,accounting for a polymorphism rate of 62.79%.The genetic similarity coefficient between the 22 phenotypic variant plants and the control group ranged from 0.5905 to 0.8286.Both the clustering analysis based on ISSR and SSR results indicated that the control group could be distinguished from the majority of the phenotypic variant plants,suggesting that most of the induced mutant plants had certain genetic differences from the control group at the molecular level.