| TRAP蛋白基因的克隆及在大肠杆菌中的高效表达 |
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| 引用本文： | 杨光,刘玉,李少华,柳川,沈倍奋,邵宁生. TRAP蛋白基因的克隆及在大肠杆菌中的高效表达[J]. 军事医学科学院院刊, 2003, 27(2): 92-95 |
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| 作者姓名： | 杨光 刘玉 李少华 柳川 沈倍奋 邵宁生 |
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| 作者单位： | 1. 军事医学科学院基础医学研究所,北京,100850
2. 白求恩军医学院,石家庄,050000 |
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| 摘 要： | 目的：研究金黄色葡萄球菌(金葡菌)毒素的调控机制，构建pET—trap表达载体，在大肠杆菌中表达金葡菌毒素调控的重要分子TRAP蛋白。方法：从金葡菌中提取基因组DNA，用特异引物钓取trap基因。通过亚克隆的方法构建了pET—trap表达载体，在大肠杆菌中诱导表达，经亲和柱层析分离纯化。制备纯化蛋白的多抗血清，利用Western印迹检测多抗与天然蛋白的反应。结果：目的基因在大肠杆菌中获得高表达，超声破碎后，表达产物主要存在于上清中，制备的多抗血清可以与天然蛋白发生特异性反应。结论：通过原核表达获得了天然的TRAP蛋白，为研究金葡菌毒素调控机制及防治金葡菌感染奠定了基础。
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| 关 键 词： | 金黄色葡萄球菌 毒素 大肠杆菌 基因克隆 基因表达 |
| 文章编号： | 1000-5501(2003)02-0092-04 |
| 修稿时间： | 2002-08-22 |
Cloning and expression of TRAP in E.coli |
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| YANG Guang ,LIU Yu ,LI Shao_Hua ,LIU Chuan ,SHEN Bei_Fen ,SHAO Ning_Sheng. Cloning and expression of TRAP in E.coli[J]. Bulletin of the Academy of Military Medical Sciences, 2003, 27(2): 92-95 |
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| Authors: | YANG Guang LIU Yu LI Shao_Hua LIU Chuan SHEN Bei_Fen SHAO Ning_Sheng |
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| Affiliation: | YANG Guang 1,LIU Yu 2,LI Shao_Hua 1,LIU Chuan 1,SHEN Bei_Fen 1,SHAO Ning_Sheng 1 |
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| Abstract: | Objective: To study the regulation of virulence factors secreted by Staphylococcus aureus , and to acquire and express trap gene in E.coli . Methods:The trap gene was acquired by PCR amplification from S.aureus genome with specific primers.The expression vector pET_trap was constructed and expressed in E.coli . The TRAP protein was purified by affinity chromatography. The polyclonal antibodies against TRAP protein were prepared and tested by Western Blot. Results: The TRAP protein was highly expressed in E.coli in soluble state. The prepared antibody can specifically recognize the natural TRAP protein. Conclusion:The acquirement of purified TRAP protein will lay a foundation for the study of the regulation of virulence factors in S.aureus . |
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| Keywords: | Staphylococcus aureus TRAP prokaryotic expression Escherichia coli |
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