唐古特大黄ISSR-PCR反应条件的优化

胡延萍<sup>1; 2</sup>; 谢小龙<sup>1; 2</sup>; 王 莉<sup>1</sup>; 杨 建<sup>1; 2</sup>; 李 毅<sup>1*</sup>

引用本文:	胡延萍, 谢小龙, 王莉, 杨建, 李毅.唐古特大黄ISSR-PCR反应条件的优化[J].广西植物,2010,(1):112-116.[点击复制]
	HU Yan-Ping, XIE Xiao-Long, WANG Li, YANG Jian, LI Yi.Establishment and optimization of ISSR-PCR reaction conditions for Rheum tanguticum[J].Guihaia,2010,(1):112-116.[点击复制]

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唐古特大黄ISSR-PCR反应条件的优化
胡延萍^1,2, 谢小龙^1,2, 王莉¹, 杨建^1,2, 李毅^1*
1.中国科学院西北高原生物研究所高原适应与进化重点实验室, 西宁 810001;2.中国科学院研究生院, 北京 100049

摘要:

利用单因素试验对影响唐古特大黄ISSR-PCR扩增的重要参数进行优化,以期建立其最佳反应条件。结果如下:20 μL反应体系包括1.5×PCR buffer(15 mmol/L Tris-HCl,75 mmol/L KCl),1.00 mmol/L MgCl₂,0.6U Taq DNA聚合酶,0.125 mmol/L dNTP,0.5 μmol/L引物和30 ng模板DNA; 引物UBC888适宜的退火温度为57.4 ℃。ISSR反应条件的建立为利用分子标记技术研究唐古特大黄居群遗传多样性奠定了良好基础。

关键词: 唐古特大黄 ISSR-PCR 优化

DOI：

分类号:Q943

基金项目:

Establishment and optimization of ISSR-PCR reaction conditions for Rheum tanguticum

HU Yan-Ping^1,2, XIE Xiao-Long^1,2, WANG Li¹, YANG Jian^1,2, LI Yi^1*

1.Key Laboratory of Adaptation and Evolution of Plateau Biota, Northwest Institute of Plateau Biology, Chinese Academy of Sciences, Xining 810001, China;2.Graduate University of Chinese Academy of Sciences, Beijing 100049, China 1.Key Laboratory of Adaptation and Evolution of Plateau Biota, Northwest Institute of Plateau Biology, Chinese Academy of Sciences, Xining 810001, China; 2.Graduate University of Chinese Academy of Sciences, Beijing 100049, China

Abstract:

Inter-Simple Sequence Repeats(ISSR)is a good molecular marker for revealing genetic diversity. Reaction system differed in different species,so optimization of ISSR-PCR reaction is very important. Factors which affect the ISSR-PCR amplification,such as the concentration of Mg²⁺,Taq DNA polymerase,dNTP,primer and template DNA with different annealing temperatures,were optimized and selected by using the genomic DNA of Rheum tanguticum as material. Optimal PCR(20 μL)mix contained 1.5×PCR buffer(15 mmol/L Tris-HCl,75 mmol/L KCl),1.00 mmol/L MgCl₂,0.6 U Taq DNA polymerase,0.125 mmol/L dNTP,0.5 μmol/L primer and 30ng template DNA. The suitable annealing temperature was 57.4 ℃ for primer UBC888. Establishment of the PCR reaction conditions could favor further studies on the genetic diversity of R.tanguticum by using ISSR molecular marker techniques.

Key words: Rheum tanguticum ISSR-PCR optimization