基于ISSR和AFLP标记开发甜菜SSR引物  

A Method for Developing Sugar Beet (Beta vulgaris L.) Microsatellite Markers Based on AFLP and ISSR Markers

作　　者：牛泽如[1] 杨文柱[1] 庞磊[1] 田自华[1] 邵金旺[1] 史树德[1] 

机构地区：[1]内蒙古农业大学农学院,呼和浩特010019

出　　处：《中国农学通报》2010年第21期147-151,共5页Chinese Agricultural Science Bulletin

基　　金：内蒙古自治区高等学校科学研究项目(NJzy08045);内蒙古农业大学博士科研启动基金;现代农业产业技术体系建设专项资金

摘　　要：基于AFLP和ISSR标记原理,采用基因组步移技术,建立了一种新的分离甜菜基因组微卫星引物的方法。具体步骤为:(1)构建基因组DNA限制性内切酶文库,ISSR引物扩增文库中两端含微卫星序列的片段并进行克隆测序,根据测序结果设计微卫星序列间的引物IP1和IP2;(2)巢式PCR进行基因组步移,扩增IP2引物下游序列并测序,进而设计微卫星引物IP3,引物IP2和IP3即为SSR标记引物。对获得的SSR引物进行PCR验证,结果表明,SSR引物产率为16%,研究获得的SSR引物具有较高的多态性,对于后续的遗传多样性检测和遗传连锁图构建具有重要意义。Based on the principle and characteristics of the molecular marker techniques of amplified AFLP and ISSR,used genome walking technology,a reliable and valid method was established for isolating microsatellites from genomic DNA of sugar beet （Beta vulgaris L.）.Fragments flanked by one or two microsatellites were amplified using a microsatellite primer,（AG） 10 or （AT） 10 etc..A primer IP l designed from the sequenced region at one end of the microsatellite,and for nested PCR another primer IP 2 based on the sequence between IP 1 and the microsatellite were prepared.Then primer IP 3 of the other side of the microsatellite DNA was designed by nested PCR.IP2/IP3 was examined as a potential SSR marker.The results showed that the success rate was 16%,which was significantly higher than that of traditional methods.The SSR primers which were got from this study showing higher polymorphic,it is great significance for the following testing of the genetic diversity and genetic linkage map construction.

关 键 词：甜菜 微卫星引物 ISSR-PCR ALFP-PCR 

分 类 号：S566.3[农业科学—作物学]