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Plant Diversity ›› 2010, Vol. 32 ›› Issue (6): 535-538.DOI: 10.3724/SP.J.1143.2010.10069

• 研究论文 • 上一篇    下一篇

一种新的用于提高lea3基因扩增特异性和产量的增强剂

于丽霞,武晓璐,汤晓倩,鄢波   

  1. 西南林业大学园林学院,云南 昆明650224
  • 收稿日期:2010-04-01 出版日期:2010-12-25 发布日期:2010-07-02
  • 基金资助:

    The Applied Foundation Research of Yunnan Province in China (Project number 2009ZC079M) and Yunnan Province and Minister Key Subject, University Key Lab and Share Platform

Effect of New Specificity and Yield Enhancer on Amplification of lea 3 Gene

 XU  Li-Xia, WU  Xiao-Lu, TANG  Xiao-Qian, YAN  Bo   

  1. Faculty of Landscape Architecture, Southwest Forestry University, Kunming 650224, China
  • Received:2010-04-01 Online:2010-12-25 Published:2010-07-02
  • Supported by:

    The Applied Foundation Research of Yunnan Province in China (Project number 2009ZC079M) and Yunnan Province and Minister Key Subject, University Key Lab and Share Platform

摘要:

在对目的DNA序列尤其是高GC含量的片段进行PCR扩增的时候,经常需要对一些试验条件进行优化。在一定条件下,二甲基亚砜、甲酰胺、甘油、NP40和Tween20等可以在某种程度上提高PCR的特异性和效率。我们在对禾本科lea3基因进行分离克隆时发现了一种新的可以提高PCR产量和特异性的物质——极高热稳定单链结合蛋白(ET SSB),研究发现在每50 μl PCR反应体系中加入200 ng的ET SSB,可以有效地抑制DNA片段的非特异性条带的产生,并可以提高目的片段的产量。

关键词: 高GC含量, 聚合酶链式反应, 极高热稳定单链结合蛋白

Abstract:

Amplification of a target DNA by the polymerase chain reaction (PCR) often requires experimental optimization factors especially when the target sequence contains high content of GC. Dimethyl sulfoxide, formamide, glycerol, NP40 and Tween20 are known to increase, the specificity and efficiency of PCR under certain conditions. In the amplification of lea3 gene in Gramineae, we found a new Extreme Thermostable SingleStranded DNA Binding Protein (ET SSB) and improved yield of PCR. Addition of 200 ng of ET SSB per 50 μl reation decreased the formation of nonspecific DNA fragments and increased the yield of specific PCR products.

Key words: High GC content, PCR, ET SSB

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