[1]沙建平,邹全明,张卫军,等.肠出血性大肠杆菌O157∶H7 LexA蛋白的表达、纯化及鉴定[J].第三军医大学学报,2008,30(12):1114-1117.
 SHA Jian-ping,ZOU Quan-ming,ZHANG Wei-jun,et al.Expression, purification and identification of EHEC O157∶H7 LexA protein[J].J Third Mil Med Univ,2008,30(12):1114-1117.
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肠出血性大肠杆菌O157∶H7 LexA蛋白的表达、纯化及鉴定(/HTML )
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《第三军医大学学报》[ISSN:1000-5404/CN:51-1095/R]

卷:
30卷
期数:
2008年第12期
页码:
1114-1117
栏目:
论著
出版日期:
2008-06-30

文章信息/Info

Title:
Expression, purification and identification of EHEC O157∶H7 LexA protein
作者:
沙建平邹全明张卫军杨珺毛旭虎郭刚罗萍刘开云陈洪章
第三军医大学医学检验系临床微生物学及免疫学教研室,重庆市生物制药工程技术研究中心
Author(s):
SHA Jian-ping ZOU Quan-ming ZHANG Wei-jun YANG Jun MAO Xu-hu GUO Gang LUO Ping LIU Kai-yun CHEN Hong-zhang
Department of Clinical Microbiology and Immunology, College of Pharmacy and Laboratory Medicine, Third Military Medical University, Chongqing 400038, China
关键词:
肠出血性大肠杆菌O157lexA纯化免疫活性
Keywords:
enterohemorrhagic Escherichia coli (EHEC) O157∶H7 purification LexA immunological activity
分类号:
R341;R378.21;R392-33
文献标志码:
A
摘要:
目的  对EHEC O157∶H7 LexA进行表达、纯化,并检测其免疫活性。  方法  lexA基因片段插入表达载体pET22b(+),在E.coli BL21(DE3)中表达。包涵体经洗涤并用8 mol/L尿素溶解,Heparin Agrose亲合柱层析为第一步纯化,Superdex75凝胶过滤层析作为第二步精细纯化,HPLC测定LexA蛋白的浓度,将纯化的LexA蛋白经注射途径免疫家兔,制备兔抗lexA血清,采用免疫双扩、ELISA及Western blot分析LexA的免疫活性。  结果  lexA以包涵体形式表达,表达产量高达总菌体蛋白的40%左右,经Heparin Agrose亲合柱层析和凝胶过滤层析两步组合纯化目的蛋白,经HPLC测定目的蛋白的最终纯度为98%,表达及纯化的lexA具有良好的免疫活性。  结论 E.coli BL21(DE3)中表达的LexA蛋白具有良好的免疫活性。
Abstract:
Objective    To clone lexA gene from enterohemorrhagic Escherichia coli (EHEC) O157∶H7, and express and purify it, and measure its immunogenicity.      Methods    The objective fragment was amplified from standard EHEC O157∶H7 cells by PCR, and then linked with the vector pET22b(+). The objective protein was induced to express in the BL21(DE3) cells after transfection. The expressed protein was purified by 2 steps of Heparin Agrose chelate affinity chromatography and gel filtration chromatography respectively. The purified lexA protein was used to immune the rabbits to prepare the polyclonal antibody. ELISA and Western blotting was used to detect the obtained serum.      Results    The expressed protein was found in insoluble form, accounted for 40% of the total bacteria protein. The final purity was 98%, which was determined by HPLC. The obtained protein was confirmed to have sound immunogenicity.     Conclusion    Our obtained recombinant LexA protein expressed in E. coli BL21(DE3) has good immunological activity.

参考文献/References:

沙建平,邹全明,张卫军,等.   肠出血性大肠杆菌O157∶H7 LexA蛋白的表达、纯化及鉴定[J]. 第三军医大学学报,2008,30(12):1114-1117.

更新日期/Last Update: 2008-06-13