布鲁菌抗原蛋白的原核表达、纯化与鉴定

发布时间：2018-03-19 17:57

本文选题：布鲁菌　切入点：原核表达　出处：《扬州大学学报(农业与生命科学版)》2017年02期 　论文类型：期刊论文

【摘要】：PCR扩增布鲁菌rplL、groEL、bp26、omp25、p39、sod1基因,构建pET-28a(+)-rplL、pCold I-groEL、pCold I-bp26、pCold I-omp25、pCold I-p39、pCold I-sod1原核表达质粒,分别转化大肠埃希菌BL21(DE3)感受态细胞,经IPTG诱导后,SDS-PAGE和Western blot分析蛋白的表达情况,免疫亲和纯化目的蛋白。结果表明:PCR分别扩增出375、1 641、753、642、1 206和522bp的目的片段,重组菌诱导后分别表达大小为20、65、33、28、47和24ku的重组蛋白,与目的蛋白理论大小相符。Western blot结果显示6种重组蛋白均能与His单抗发生特异性反应,表明6种重组蛋白具有良好的免疫反应性,为进一步深入研究奠定了基础。
[Abstract]:The pET-28a (pET-28a) pET-28a was amplified by PCR. The prokaryotic expression plasmid of pET-28a was constructed by SDS-PAGE and Western blot analysis, and the expression of pET-28a was analyzed by SDS-PAGE and Western blot analysis after SDS-PAGE and Western blot were used to analyze the expression of pET-28a, and the expression of the protein was analyzed by SDS-PAGE and Western blot, respectively, and the expression of pET-28a was analyzed by SDS-PAGE and Western blot, respectively, and the expression of pET-28a was analyzed by SDS-PAGE and Western blot, respectively. The expression of pET-28a was analyzed by SDS-PAGE and Western blot. The purified target protein was purified by immuno-affinity assay. The results showed that the target fragments of 375U 1 641C 753N 642N 1 206 and 522bp were amplified by PCR, respectively, and the recombinant proteins were expressed at the size of 20x65m3328447 and 24ku, respectively, after induction by the recombinant bacteria. The results of Western blot showed that the six recombinant proteins could react specifically with His McAbs, indicating that the six recombinant proteins had good immunoreactivity, which laid a foundation for further study.
【作者单位】：扬州大学江苏省人兽共患病学重点实验室/江苏省动物重要疫病与人兽共患病防控协同创新中心;
【基金】：国家自然科学基金资助项目(31602031) 江苏省重点研发计划项目(BE2015343) 江苏省自然科学基金资助项目(BK20160466) 江苏高校青蓝工程项目(2016-05) 江苏高校优势学科建设工程项目(2014-05)
【分类号】：R378.5

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