玉米品种真实性和纯度鉴定的SSR标记多重PCR体系优化  

Optimization on multiplex PCR amplification system of SSR marker for authenticity and purity identification of maize varieties

作　　者：常宏[1,2] 王汉宁[1,3] 张金文[1,3] 王威[1] 路则权[1] 李瑛[1] 王蒂[1,3] 

机构地区：[1]甘肃农业大学农学院,甘肃兰州730070 [2]甘肃省种子管理总站,甘肃兰州730020 [3]甘肃省作物遗传改良与种质创新重点实验室,甘肃兰州730070

出　　处：《草业学报》2010年第2期204-211,共8页Acta Prataculturae Sinica

基　　金：国家高新技术研究发展计划(863计划)项目(2004AA207140)资助

摘　　要：为了建立稳定可靠的玉米真实性和纯度鉴定SSR标记,对DNA提取方法、SSR引物和多重PCR反应程序进行了优化。结果表明,用预热到75℃以上的研钵和95℃的1.5×CTAB提取缓冲液进行材料研磨,可得到纯度高、完整性好的DNA,并且提取成本较低。利用软件Primer Premier 5.0和Oligo 6.72对玉米指纹鉴定的SSR核心引物进行重新分析与设计,建立了21对SSR通用引物构成的8组多重PCR复合扩增体系和3步法扩增程序,均能在统一的PCR扩增条件下进行,扩增片段之间不存在交叉现象,扩增条带清晰,扩增结果稳定,这一扩增体系检测效率比单对SSR引物提高2.6倍以上。In order to establish a stable and reliable identification of maize authenticity and purity of the SSR markers,genetic DNA extraction,SSR primers and multiplex PCR reaction procedures were optimized.The results showed that the high purity and good integrity of DNA could be obtained using above 75℃ preheating mortar and 95℃ of 1.5×CTAB extraction buffer to grind material.Redesign and optimize the primer sequence using the software Primer Premier 5.0 and Oligo 6.72,eight sets of multiplex PCR amplified reactions,which was consisting of 21 pairs of universal SSR primers and a three-step procedure,were with uniform procedures,no intersection among the amplified fragments,amplified bands clear and stable amplification results.The testing efficiency of the amplification system increased 2.6 times higher than a single pair of primers SSR.

关 键 词：玉米 真实性 纯度 DNA提取 SSR引物 多重PCR 

分 类 号：S513.01[农业科学—作物学]