摘要：
以紫花苜蓿(Medicago sativa)不育系MS-GN-1 A为母本,恢复系MS178 为父本组合构建BSA分离群体,获得的F1代均表现为雄性可育,在大田种植了221株 F2代群体单株,盛花期将花粉颗粒染色后,显微镜下观察统计出,不育的F2代株数为57,可育F2代株数164,并没有观察到半不育植株.将所有植株划分为可育和不育两组,并构建可育和不育DNA混池,混池DNA从可育和不育植株组DNA中各随机抽取20个样品,以此对恢复基因定位.随机挑选160对已知的四倍体苜蓿 SSR引物扩增基因池DNA,获得2个具有多态性的分子标记,分别是 Mt2c12、AW166,初步定位Rf 基因在类群Composite5 上.将类群Composite5 上的所有引物进行合成,进一步进行引物筛选,最终获得 4 个具有多态性的标记BI68、Mt2c12、BG267和 AW776153,遗传距离分别为19.0、20.9、44.6和72.1 cM.%In this study,alfalfa CMS MS-GN-1A was used as the female parent,and the restorer line MS 178 was used as the male parent to construct BSA separate groups.All products of F1 were male fertile.There were 221 plants in the F2,and the number of sterile plants was 57,with 164 fertile,and no semi-sterile plants were found.Based on the results of the fertility microscopy,the separate population was divided into two groups, fertile and sterile group;the genomic DNA of the fertile and sterile groups were extracted and 20 samples of DNA from each group were randomly selected and mixed into a fertile and sterile gene pool for the positioning of restoring genes.The experiment randomly selected 160 pairs of known tetraploid alfalfa SSR primers.To amplify the gene pool DNA,two molecular markers with polymorphism(Mt2c12 and AW166)were obtained, and were initially positioned on a composite of five primers.All primers on this group were synthesized,and the designed primers were screened again.Finally,four markers with polymorphism were obtained:BI68,Mt2c12, BG267 and AW776153,with a genetic distance of 19.0 cM,20.9 cM,44.6 cM,and 72.1 cM,respectively.
