›› 2007, Vol. 15 ›› Issue (5): 429-436.DOI: 10.11733/j.issn.1007-0435.2007.05.006

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Optimization of SSR Reaction System of Medicago truncatula and Its Application in Annual Medicago Germplasm Identification

ZHANG Li-fang, WEI Zhen-wu, YANG Zhan-hua   

  1. Pratacultural College, Gansu Agricultural University, Lanzhou, Gansu Province 730070, China
  • Received:2007-04-05 Revised:2007-06-13 Online:2007-10-15 Published:2007-10-15

蒺藜苜蓿SSR反应体系优化及在一年生苜蓿种质鉴定中的应用

张丽芳, 魏臻武, 杨占花   

  1. 甘肃农业大学草业学院, 兰州, 730070
  • 通讯作者: 魏臻武,E-m ail:zh enw u_w ei@yahoo.com.cn
  • 作者简介:张丽芳(1981- ),女,汉族,甘肃定西人,硕士研究生,研究方向为牧草种质资源评价和育种,E-mail:zhanglifang1981@163.com
  • 基金资助:
    甘肃省教育厅科学技术研究项目(0507);甘肃农业大学科技创新项目(200518);草业生态系统教育部省部共建重点实验室和国家自然科学基金(30671486)

Abstract: SSR reaction conditions on annual medics were optimized. The model legume, Medicago truncatula., was used to set up a SSR-PCR amplification system for the annual medics. The results show that the optimal SSR-PCR reaction system for annual medics was 1 U TaqDNA polymerase, 0.20 mmol/L dNTP, 2.0 mmol/L Mg2+, 1.5 μmol/L primer, and 20 ng/μL DNA in the total volume of 10 μL. Through above PCR system, SSR fragments of 19 annual medics’ germplasms were obtained and detected. Polymorphism between different lines was abundantly detected by 8% polyacrylamide gel. Germplasms of 19 annual medics were definitely differentiated using cluster analysis based on the UPMGA.

Key words: Annual medic, Medicago truncatula Gaertn., SSR reaction, Orthogonal design, Hybridization identification, Germplasm identification

摘要: 利用正交设计和完全随机试验优化蒺藜苜蓿(Medicago truncatula Gaertn)SSR标记的PCR反应体系。结果表明,适合于一年生苜蓿(Medicago L.)遗传分析的SSR技术体系为:10μL反应体系中TaqDNA聚合酶为1U,dNTP浓度为0.20mmol/L,Mg2+浓度为2.0mmol/L,引物浓度为1.5umol/L,模板DNA用量为20ng/uL;利用该反应体系将2个蒺藜苜蓿亲本和杂交种有效鉴别;利用SSR标记对19个一年生苜蓿种质进行SSR-PCR扩增,用8%的非变性聚丙烯酰胺凝胶电泳检测,不同种质间DNA谱带多态性丰富,通过UPGMA方法聚类分析可以明确区分19个一年生苜蓿种质。

关键词: 一年生苜蓿, 蒺藜苜蓿, SSR反应, 正交设计, 杂交种鉴定, 种质鉴定

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