| 霍乱毒素B亚单位基因的克隆和表达 |
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| 引用本文： | 田季德 徐兆炜. 霍乱毒素B亚单位基因的克隆和表达[J]. 中国医学科学院学报, 1991, 13(5): 332-337 |
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| 作者姓名： | 田季德 徐兆炜 |
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| 作者单位： | 中国预防医学科学院流行病学微生物学研究所 北京(田季德，徐兆炜，高庆申)，中国预防医学科学院流行病学微生物学研究所 北京(刘延清) |
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| 摘 要： | 用DNA重组技术构建CT-B表达性质粒pXB1及进一步修饰的pXB2,使CT-B基因在大肠杆菌中得到较高水平的表达(280～350ng/ml),且有71.5％分泌率。表达动力学研究阐明,克隆子培养24h产物收率较高。一定浓度的Mg~(2+)、L-组氨酸和天冬酰胺能刺激CT-B的表达。GM1-ELISA、CHO细胞测毒结合间接免疫荧光检测和家兔肠段结扎试验证实,表达的CT-B能与游离的或活细胞膜上的GM1受体结合,但无细胞和肠段毒性。这种细胞测毒结合免疫荧光序贯试验,为客观证实CT-B的生物学活性开拓了新途径。
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| 关 键 词： | 霍乱毒素 B亚单位 基因克隆 |
Cloning and Expression of the Cholera Toxin B Subunit Gene |
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| Tian Jide,. Cloning and Expression of the Cholera Toxin B Subunit Gene[J]. Acta Academiae Medicinae Sinicae, 1991, 13(5): 332-337 |
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| Authors: | Tian Jide |
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| Affiliation: | Institute of Epidemiology and Microbiology, Chinese Academy of Preventive Medicine, Beijing. |
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| Abstract: | The CT-B gene-expressing plasmid pXB1 and modified pXB2 were constructed using recombinant DNA techniques. These resulted in high CT-B gene expression in E. coli (280-350 ng/ml), with CT-B accounting for 71.5% of total extracellular protein. The results of CT-B expression kinetics showed that the CT-B product levels reached a peak after 24 h in culture. Certain concentrations (100 micrograms/ml) of MgCl2, L-histidine and asparagine can increase the CT-B product level by 10%-25%. The binding of the expressed CT-B product to GM1 receptor on free or live cell membranes was identified by GM1-ELISA, CHO cytotoxicity test, indirect immunofluorescence, and the rabbit ileal loop test. No toxic response to cells or rabbits was detected. |
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| Keywords: | cholera toxin B subunit gene clone gene expression |
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