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| 霍乱弧菌zot基因的克隆及其在大肠杆菌中的表达 | |
| 引用本文： | 何志勇,陈哲宇,王东宁,杨冠珍,张惟杰,吴祥甫.霍乱弧菌zot基因的克隆及其在大肠杆菌中的表达[J].生物工程学报,2000,16(5):570-573. |
| 作者姓名： | 何志勇 陈哲宇 王东宁 杨冠珍 张惟杰 吴祥甫 |
| 作者单位： | 1. 中国科学院上海生物化学研究所,上海,200031;上海交通大学生物科学与技术系,上海,200240 2. 第二军医大学神经生物学教研室 3. 中国科学院上海生物化学研究所,上海,200031 4. 上海交通大学生物科学与技术系,上海,200240 |
| 摘 要： | 从霍乱疫苗菌中抽提基因组ＤＮＡ，用ＰＣＲ的方法扩增ｚｏｔ基因。序列分析表明，ｚｏｔ基因编码３９９个氨基酸，其中４个氨基酸与文献报道有差异。将ｚｏｔ基因插入含Ｔ７启动子的质粒ｐＥＴ－２８（ａ＋）构建表达质粒ｐＥＴ－ＺＯＴ，转化大肠檑菌ＢＬ２１（ＤＥ３）筛有达菌株ＢＬＺＯＴ。表达菌株经１ｍｍｏｌ／ＬＴ ＩＰＴＧ诱导表达３－５ｈ后，表达大量ＺＯＴ蛋白，并形成包涵体，经ＳＤＳ－ＰＡＧＥ分析重组ＺＯＴ蛋白分 |
| 关 键 词： | 封闭带毒素 霍乱弧菌 基因药物 口服途径 |
| 文章编号： | 1000-3061(2000)05-0570-04 |
| 修稿时间： | 1999-11-22 |
Cloning of the zot Gene of Vibrio cholerae and Its Expression in Escherichia coil |
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| HE Zhi-Yong,CHEN Zhe-Yu,WANG Dong-Ning,YANG Guan-Zhen,ZHANG Wei-Jie,WU Xiang-Fu.Cloning of the zot Gene of Vibrio cholerae and Its Expression in Escherichia coil[J].Chinese Journal of Biotechnology,2000,16(5):570-573. | |
| Authors: | HE Zhi-Yong CHEN Zhe-Yu WANG Dong-Ning YANG Guan-Zhen ZHANG Wei-Jie WU Xiang-Fu |
| Affiliation: | HE Zhi-Yong ;(Shanghai Institute of Biochemistry, Chinese Academy of Sciences, Shanghai 200031 Department of Bioscience and Biotechnology, Shanghai Jiao-Tong University, Shanghai 200240);CHEN Zhe-Yu ;(Shanghai Institute of Biochemistry, Chinese Academy of Sciences, Shanghai 200031);WANG Dong-Ning ;(Shanghai Institute of Biochemistry, Chinese Academy of Sciences, Shanghai 200031 Department of Bioscience and Biotechnology, Shanghai Jiao-Tong University, Shanghai 200240);YANG Guan-Zhen ;(Shanghai Institute of Biochemistry, Chinese Academy of Sciences, Shanghai 200031);ZHANG Wei-Jie ;(Department of Bioscience and Biotechnology, Shanghai Jiao-Tong University, Shanghai 200240);WU Xiang-Fu ;(Shanghai Institute of Biochemistry, Chinese Academy of Sciences, Shanghai 200031) |
| Abstract: | The zot gene encoding Zonula occludens toxin was amplified from classic Vibrio cholerae genomic DNA by PCR.The result of sequencing indicated that zot gene encodes 399 amino acid residues.The sequence of zot gene was a little bit different from that of reported including 14 nucleotides and four amino acid residues.The expression plasmid pET\|ZOT was constructed by inserting zot gene into plasmid pET\|28a( ) containing the T7 promoter.The expression plasmid was induced into E.coli BL21 (DE3) and expression strain BLZOT was selected.SDS PAGE analysis revealed that the ZOT protein was expressed and accumulated up to above 15% of bacterial soluble protein after induced by IPTG.A protein of 47kD was expressed as including body.Western blot analysis revealed that the expressed protein was ZOT. |
| Keywords: | Zonula occludens toxin gene cloning gene expression E coli |
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