利用SCoT标记分析不同秋眠型苜蓿的遗传多样性  

Optimization of SCoT reaction system and genetic diversity of different fall dormancy alfalfa

作　　者：何庆元[1,2] 王吴斌[2] 杨红燕[2] 向仕华[2] 周丽英[1] 王松华[1] 

机构地区：[1]安徽科技学院生命科学学院,安徽凤阳233100 [2]南京农业大学大豆研究所国家大豆改良中心作物遗传与种质创新国家重点实验室,江苏南京210095

出　　处：《草业学报》2012年第2期133-140,共8页Acta Prataculturae Sinica

基　　金：国家"973"计划基金项目(2007CB108901);安徽省教育厅重大基金项目(ZD200910);安徽科技学院校级重点学科(AKXK20102-1)资助

摘　　要：用L16(45)正交实验设计研究模板DNA、dNTPs、Mg2+、Taq酶和引物5个因素的浓度对SCoT扩增迁移率重现率的影响。结果表明,在20μL反应体系中,2.5ng/μL DNA、1.00mmol/L Mg2+、1.20U Taq酶、0.40mmol/L dNTPs和0.30μmol/L浓度的引物最为稳定,重现率达到100%。利用最优反应体系从50条引物中筛选出13条扩增效果好的引物,将它们分别扩增34个苜蓿品种,共检测到103个SCoT标记,其中92个位点具有多态性。聚类分析表明,从安徽和江苏两地收集的野生南苜蓿和栽培苜蓿的遗传距离最远,单独聚为一类;其余32个品种苜蓿在相似系数0.757附近分为3个亚群,秋眠型苜蓿品种主要分布在第Ⅱ亚群中,半秋眠型在3个亚群中都有分布,而非秋眠型苜蓿分布在第一和第二亚群中,表明SCoT标记的聚类结果在一定程度上能够反映苜蓿的秋眠型,但并不完全一致。Genomic DNA was extracted by advanced CTAB methods from Medicago sativa and Medicago hispi- da. The optimum SCoT--PCR （start eodon targeted polymorphism） reaction system in alfalfa was established with orthogonal design experiments on four levels of five factors （DNA, dNTPs, Mg2＋ , Taq poIymerase, and primer）. A suitable SCoT reaction system was a 20 μL mixture containing 50 ng DNA, 1.00 mmol/L Mg2＋ , 1.20 U Taq polymerase, 0.40 mmol/L dNTPs and 0.30μmol/L primers. Of the 50 primers screened, 13 gen- erated highly polymorphic fragments useable as SCoT markers, and 34 accessions were amplified by these 13 primers. A total of 103 bands （including 92 po!ymorphic bands） were detected by the 13 chosen primers. The 34 accessions were divided into two groups. Two M. hispida cultivars from Anhui and Jiangsu were clustered into one group. The remaining 32 cultivars were classified into three sub-groups with a similarity coefficient of O. 757. Fall dormancy alfalfa cultivars were mostly distributed in the second sub-group. Semi-fall dormancy and non-fall dormancy alfalfa cultivars did not converge into the same sub-group thus showing incomplete similarity between fall dormancy alfalfas.

关 键 词：苜蓿 SCoT 条件优化 秋眠型 遗传多样性 

分 类 号：S541.9[农业科学—作物学]