pPET-SUMF2各亚型的原核质粒构建及蛋白表达

发布时间：2018-06-28 22:04

本文选题：硫酸酯酶修饰因子 + 原核表达载体　；参考：《中国科技论文》2014年09期

【摘要】：构建人硫酸酯酶修饰因子2(SUMF2)基因各亚型的融合表达载体并在大肠杆菌中诱导表达pPET-SUMF2各亚型的融合蛋白。用EcoRI和XhoI双酶切人SUMF2各种亚型基因及质粒pPET-30a;将2种酶切产物按常规方法连接、转化至大肠杆菌DH5α。挑取菌落培养,提取质粒进行验证。将构建成功的pPET-SUMF2各亚型重组表达质粒转化至BL21(DE3)中,经IPTG诱导后,SDS-PAGE电泳分析以及Western blot检测SUMF2各亚型的融合蛋白表达。成功构建了pPET-SUMF2各亚型的重组表达质粒,并成功诱导SUMF2各亚型融合蛋白的表达,Western blot结果显示在预期位置有特异蛋白条带表达,并在BL21(DE3)内表达了SUMF2各亚型的融合蛋白,为进一步研究SUMF2的功能以及SUMF2与哮喘发病的关系奠定了基础。
[Abstract]:The fusion expression vector of human sulfate-modifying factor 2 (SUMF2) gene subtype was constructed and the fusion protein of pPET-SUMF2 subtype was induced and expressed in Escherichia coli. Human SUMF2 subtype genes and plasmids pPET-30awere digested with EcoRI and XhoI, and the two products were ligated into E. coli DH5 伪 by routine method. The colony culture was carried out and the plasmid was extracted for verification. The recombinant expression plasmid pPET-SUMF2 was transformed into BL21 (DE3). After induced by IPTG, SDS-PAGE electrophoresis and Western blot were used to detect the expression of fusion protein of SUMF2 subtype. The recombinant expression plasmid of each subtype of pPET-SUMF2 was successfully constructed, and the expression of subtype fusion protein of SUMF2 was induced successfully. The results of Western blot showed that there were specific protein bands in the expected position, and the fusion protein of each subtype of SUMF2 was expressed in BL21 (DE3). It lays a foundation for further study of the function of SUMF2 and the relationship between SUMF2 and asthma.
【作者单位】：哈尔滨医科大学附属第四医院检验科;
【基金】：高等学校博士学科点专项科研基金资助项目(20112307110021) 国家自然科学基金资助项目(81171657)
【分类号】：R562.25

【共引文献】