| 羊布鲁菌外膜蛋白Omp22的原核表达及鉴定 |
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| 引用本文： | 张亮,张蕾,张芳琳,李璞媛,李凯,吴兴安,徐志凯,白文涛.羊布鲁菌外膜蛋白Omp22的原核表达及鉴定[J].科学技术与工程,2009,9(14). |
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| 作者姓名： | 张亮 张蕾 张芳琳 李璞媛 李凯 吴兴安 徐志凯 白文涛 |
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| 作者单位： | 第四军医大学微生物学教研室,西安,710032 |
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| 基金项目： | 军队科技攻关项目(06G091);;第四军医大学科研重点项目(05XJZ006)资助 |
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| 摘 要： | 布鲁菌外膜蛋白Omp22与布鲁菌致病性相关,又可诱导机体免疫反应。为进一步研究Omp22的特性,对其进行原核表达,并初步鉴定。根据Genbank序列,设计并合成羊布鲁菌外膜蛋白Omp22引物。以羊布鲁菌M5株基因组DAN为模板,PCR扩增并克隆入原核表达载体pGEX-4T-1中。酶切鉴定正确后,测定序列;将重组质粒pGEX-4T-1-Omp22电转化大肠杆菌DH5α中,诱导表达,并采用羊布鲁菌兔免疫血清,Western-blot初步检测融合蛋白GST-Omp22表达情况和免疫学特性。结果PCR扩增出约660bp目的条带,与预期相符;酶切鉴定和测序结果表明,成功构建了pGEX-4T-1-Omp22原核表达载体;优化诱导温度,结果表明,在25℃诱导表达时,该融合蛋白大部分出现在上清中;Western-blot结果证实,GST-Omp22可与羊布鲁菌兔免疫血清特异性结合。说明成功构建了Omp22蛋白原核表达载体并进行了可溶性表达;该融合蛋白可与羊布鲁菌兔免疫血清特异性结合。Omp22蛋白的表达,为进一步研究布鲁菌的致病机制以及疫苗研制奠定基础。
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| 关 键 词： | 羊布鲁菌 外膜蛋白Omp22 原核表达 鉴定 |
Prokaryotic Expression and Identification of Omp22 Protein of Brucella melitensis |
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| ZHANG Liang,ZHANG Lei,ZHANG Fang-lin,LI Pu-yuan,LI Kai,WU Xing-an,Xu Zhi-kai,BAI Wen-tao.Prokaryotic Expression and Identification of Omp22 Protein of Brucella melitensis[J].Science Technology and Engineering,2009,9(14). |
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| Authors: | ZHANG Liang ZHANG Lei ZHANG Fang-lin LI Pu-yuan LI Kai WU Xing-an Xu Zhi-kai BAI Wen-tao |
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| Institution: | Department of Microbiology;the Fourth Military Medical University;Xi'an 710032;P.R.China |
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| Abstract: | Omp22 is involved in virulence in Brucella melitensis and its immunogenesis.To clone and express the brucella outer membrane protein Omp22,the gene of the protein was amplified from Brucella melitensis with PCR method,and then the PCR product was subcloned into vector pGEX-4T-1 and expressed in E.coli.The recombinant protein was detected by Western-blot.According to the SDS-PAGE and Western-blot analysis,the recombinant plasmid pGEX-4T-1-Omp22 could be expressed successfully in E.coli and the fusion protein... |
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| Keywords: | Brucella melitensis Omp22 cloning prokaryotic expression identification |
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