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| 单增李斯特氏菌actA基因的克隆及原核表达 | |
| 引用本文： | 张辉,王兴龙,王俊霞,李晓艳,卜昭阳,张爱玲,崔丽瑾,闫广谋.单增李斯特氏菌actA基因的克隆及原核表达[J].畜牧与兽医,2008,40(3):14-17. |
| 作者姓名： | 张辉 王兴龙 王俊霞 李晓艳 卜昭阳 张爱玲 崔丽瑾 闫广谋 |
| 作者单位： | 1. 吉林大学农学部畜牧兽医学院,吉林,长春,130062;军事医学科学院军事兽医研究所,吉林,长春,130062 2. 军事医学科学院军事兽医研究所,吉林,长春,130062 3. 吉林大学农学部畜牧兽医学院,吉林,长春,130062 |
| 摘 要： | 利用PCR技术从单增李斯特氏菌中扩增出actA基因,将PCR产物纯化后克隆到pMD 18Tsimple vector中,成功构建出克隆载体pMD-18T/actA。以BamHⅠ和EcoRⅠ分别双酶切pMD-18T/ActA和表达载体pGEX-3X,将纯化的actA基因亚克隆到表达载体pGEX-3X。构建的重组表达载体pGEX-3X/ActA转化到E.coli BL21(DE3)中,IPTG诱导表达,表达产物进行SDS-PAGE分析,可见约120 ku外源蛋白带,Western blot分析表明该蛋白可与单增李斯特氏菌多克隆抗体发生特异性反应。该研究为ActA的生物学特性和功能的研究及诊断试剂的研制奠定了基础。 |
| 关 键 词： | 单增李斯特氏菌 actA基因 克隆 原核表达 |
| 文章编号： | 0529-5130(2008)03-0014-04 |
| 修稿时间： | 2007年10月12 |
Cloning and prokaryotic expression of actA gene of Listeria monocytogens |
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| ZHANG Hui,WANG Xing-long,WANG Jun-xia,LI Xiao-yan,PU Zhao-yang,ZHANG Ai-ling,CUI Li-jin,YAN Guang-mou.Cloning and prokaryotic expression of actA gene of Listeria monocytogens[J].Animal Husbandry & Veterinary Medicine,2008,40(3):14-17. | |
| Authors: | ZHANG Hui WANG Xing-long WANG Jun-xia LI Xiao-yan PU Zhao-yang ZHANG Ai-ling CUI Li-jin YAN Guang-mou |
| Abstract: | The gene encoding ActA was amplified from Listeria monocytogens by PCR.Cloning vector pMD-18T/ActA was successfully constructed by cloning the PCR product into pMD 18T simple vector.pMD-18T/ActA and pGEX-3X were digested by BamHⅠ and EcoRⅠ.The recombinant expression vector pGEX-3X/ActA was constructed by subcloning the actA gene into the expression vector pGEX-3X,and transformed into E.coli BL21(DE3).The bacteria were induced by IPTG and the lysates were analyzed by SDS-PAGE.The expressed fusion protein showed a molecular weight of 120 ku.The protein could react with polyclonal antibody against L.monocytogens in Western blot.The results provide a foundation for further studies on the biological characteristics of ActA and its role in listeria pathogenesis and for diagnosis agent. |
| Keywords: | Listeria monocytogens actA gene cloning prokaryotic expression |
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