Integrative Biology Journals

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

• Articles • Previous Articles     Next Articles

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:2010-07-02 Published:2010-12-25
  • 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

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
  • 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

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

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

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