红斑丹毒丝菌C43065株spaA基因的克隆和表达 |
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引用本文: | 刘丹丹,杨振龙,吾鲁木汗·那孜尔别克.红斑丹毒丝菌C43065株spaA基因的克隆和表达[J].吉首大学学报(自然科学版),2013,34(5):85-88. |
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作者姓名: | 刘丹丹 杨振龙 吾鲁木汗·那孜尔别克 |
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作者单位: | (吉首大学生物资源与环境科学学院,湖南 吉首 416000) |
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基金项目: | 国家自然科学基金资助项目(31072142) |
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摘 要: | 通过PCR从红斑丹毒丝菌C43065株基因组DNA中扩增出编码信号肽除外的成熟SpaA蛋白基因spaA,将其克隆到表达载体pET32a的BamHⅠ和Hind Ⅲ位点上,构建重组表达质粒pET-spaA,转化大肠杆菌BL21,在IPTG诱导下表达N端带有Trx标签的融合蛋白rSpaA,SDS-PAGE检测表达蛋白.DNA测序结果表明,spaA基因大小为1794 bp,编码由597个氨基酸残基组成的成熟SpaA蛋白,SDS-PAGE结果显示在大肠杆菌BL21中成功表达了分子量约为86 kDa的重组rSpaA,为进一步开展SpaA保护区域的研究奠定基础.
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关 键 词: | 红斑丹毒丝菌 spaA基因 克隆 原核表达 |
Cloning and Expression of spaA Gene of Erysipelothrix Rhusiopathiae C43065 |
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Institution: | (College of Biology and Environmental Sciences,Jishou University,Jishou 416000,China) |
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Abstract: | The spaA gene encoding mature surface protective antigen A (SpaA) without signal peptide was amplified from genomic DNA of E. rhusiopathiae C43065 by PCR,The BamH I and HindⅢ digested PCR product was cloned into prokaryotic expression vector pET32a to generate a recombinant plasmid pET-spaA. The recombinant protein rSpaA was expressed in E. coli BL21 harboring the recombinant plasmid pET-spaA by IPTG inducing, and the expressed protein was determined by SDS-PAGE. The DNA sequence analysis showed that the spaA gene of C43065 strain was 1794 bp in length. SDS-PAGE a- nalysis revealed a single protein band with a molecular weight of 86 kDa successfully expressed in E. coli BL21. The expressed protein of rSpaA will contribute to further study on protective domain of this pro- tein. |
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Keywords: | Erysipelothrix rhusiopathiae spaA gene cloning prokaryotic expression |
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