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HindⅢ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1794bp,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86kDa的重组rSpaA,为进一步开展SpaA保护区域的研究奠定基础.
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.
出处
《吉首大学学报（自然科学版）》
CAS
2013年第5期85-88,共4页
Journal of Jishou University(Natural Sciences Edition)
基金
国家自然科学基金资助项目(31072142)
关键词
红斑丹毒丝菌
spaA基因
克隆
原核表达
Erysipelothrix rhusiopathiae spaA gene
cloning
prokaryotic expression
