MOLECULAR CLONING AND EXPRESSION OF <italic style="font-style: italic">lexA</italic> GENE FROM THE RADIORESISTANT BACTERIUM <italic style="font-style: italic">Deinococcus radiodurans</italic>

WANG Mingsuo; DU Zeji; KONG Xiangrong; SU Liaoyuan

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MOLECULAR CLONING AND EXPRESSION OF lexA GENE FROM THE RADIORESISTANT BACTERIUM Deinococcus radiodurans

更新时间：2023-04-07

- MOLECULAR CLONING AND EXPRESSION OF lexA GENE FROM THE RADIORESISTANT BACTERIUM Deinococcus radiodurans
- Journal of Radiation Research and Radiation Processing Vol. 18, Issue 3, Pages: 193-197(2000)
- 作者机构：
  
  1.苏州医学院放射医学系　苏州　215007
- 作者简介：
- 基金信息：
  
  the Special Research Fundation of Hygiene Ministry for Excellent Young Seientists and granted in the year 1999
- DOI：
  CLC：
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WANG Mingsuo, DU Zeji, KONG Xiangrong, et al. MOLECULAR CLONING AND EXPRESSION OF lexA GENE FROM THE RADIORESISTANT BACTERIUM Deinococcus radiodurans. [J]. Journal of Radiation Research and Radiation Processing 18(3):193-197(2000)
DOI：

WANG Mingsuo, DU Zeji, KONG Xiangrong, et al. MOLECULAR CLONING AND EXPRESSION OF lexA GENE FROM THE RADIORESISTANT BACTERIUM Deinococcus radiodurans. [J]. Journal of Radiation Research and Radiation Processing 18(3):193-197(2000) DOI：

摘要

研究了旨在克隆抗辐射菌,Deinococcus radiodurans,的,lexA,基因并构建其表达载体，以便进一步研究,lexA,基因的功能及其在辐射抗性中的作用。主要的实验步骤包括分离提取抗辐射菌基因组DNA，分离出,lexA,基因并测定其序列，克隆于质粒载体PUC19, 用电击穿孔法将重组的质粒导入大肠杆菌JM109, SDS-PAGE检测基因的表达。用定点突变技术，改变,lexA,基因核糖体结合位点（RBS）, 以增强,lexA,基因的表达。结果表明，,lexA,基因位于基因组DNA的,BlnI-AscI,片断中，由630个碱基对（bp）组成，编码210个氨基酸（aa）, 理论上推定，分子量为22.5kD，等电点为6.4。用pUC19构建的,lexA,基因表达载体能在大肠杆菌JM109中表达，但表达效率低，改变,lexA,的核糖体结合位点RBS可提高该基因表达水平，使进一步分离纯化,lexA,蛋白质以及深入研究该基因的作用成为可能。

Abstract

In order to investigate the role of ,lexA, gene in the radioresistance of ,Deinococcus radiodurans, the expressing vector of ,lexA, gene was constructed. The genomic DNA was extracted from wild type ,Deinococcus radiodurans, KD8301. The ,lexA, gene was isolated from the genomic DNA and sequenced. The Plasmid vector pUC19 was used to clone ,lexA, gene and the electroporation was employed to transfer the recombinant plasmid into ,E. coli, JM109. SDS-PAGE was used to check the expression of ,lexA, gene. The ribosome binding site (RBS) of ,lexA, gene was mutated by means of Site -Directed Mutagenesis technique for the optimum of gene expression. The results indicated that the ,lexA, gene was located in the ,Blnl-AscI, fragment of ,Deinocococus radiodurans, genomic DNA, containing 630bp and coding 210 aa. The theoretically deduced molecular weight and the isoelectric point were 2.5KDa and 6.4 respectively. ,lexA, gene inserted into pUC19 could be expressed in JM109, but at very low level. After optimizing the RBS of ,lexA, gene, higher level of ,lexA, expression was obeserved. The results make it possible to isolate and purify ,lexA, protein, and further to investigate the function of ,lexA, gene in ,Deinococcus radiodurans.

关键词

抗辐射菌lexA基因基因克隆基因表达

Keywords

Deinococcus radioduransLexA geneGene cloningGene expression

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MOLECULAR CLONING AND EXPRESSION OF lexA GENE FROM THE RADIORESISTANT BACTERIUM Deinococcus radiodurans

DOI：

摘要

Abstract

关键词

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