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双歧杆菌表达肠致病性大肠埃希菌EspA蛋白的体外研究

, PP. 993-994

Keywords: 肠致病性大肠埃希菌,EspA基因,双歧杆菌,蛋白诱导表达

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Abstract:

目的构建肠致病性大肠埃希菌(EnteropathogenicE.coli,EPEC)EspA蛋白双歧杆菌表达重组菌株,并验证蛋白表达。方法采用常规PCR扩增EspA片段,体外构建重组质粒pBAD-EspA,转化大肠杆菌BL21感受态细胞培养,挑取单克隆提取质粒,双酶切和PCR对重组质粒鉴定。pBAD-EspA电转化长双歧杆菌感受态细胞,挑取阳性克隆并PCR鉴定,经木聚糖诱导表达,蛋白质印迹试验(Westernblot)分析验证EspA蛋白在双歧杆菌中的表达。结果经酶切后琼脂糖凝胶电泳及测序结果证明pBAD-EspA重组质粒构建成功。pBAD-EspA重组质粒转化双歧杆菌后经阿拉伯糖诱导,在诱导表达不同时间点菌液沉淀均有良好的蛋白表达。结论体外成功构建表达EPECEspA蛋白的长双歧杆菌,为制备ESPA口服疫苗提供科学依据。

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