Sun Su-xia, Wang Hong, Wang Yong, Xiang Qian, Wu Ai-jun, Yu Shou-yi
Department of Epidemiology, First Military Medical University, Guangzhou 510515, China.
Di Yi Jun Yi Da Xue Xue Bao. 2003 Mar;23(3):230-2.
To induce the expression of and purify invasion plasmid antigen C (IpaC) of Shigella flexneri for studying the pathogenesis of Shigella flexneri.
Prokaryotic expression plasmid pET32a-ipaC was constructed and incorporated into E.coli BL21 (lambda DE3). The engineered bacteria were induced by isopropyl-beta-D-thiogalactopyranoside (IPTG) to express IpaC, which was identified by SDS-PAGE and purified by QIA expressionist system.
SDS-PAGE presented a band for the fusion protein with the relative molecular mass of approximately 63 000, whose expression reached up to 11% of the total protein of E.coli BL21(lambda DE3). After proper purification, a purity of the target fusion protein of over 90% was achieved when the concentration of imidazole for elution was 350 mmol/L.
The recombinant plasmid pET32a-ipaC has been stably and efficiently expressed in E.coli BL21 (lambda DE3), and QIA expressionist purification system proves to be simple and highly efficient.
诱导福氏志贺菌侵袭质粒抗原C（IpaC）的表达并进行纯化，以研究福氏志贺菌的致病机制。
构建原核表达质粒pET32a-ipaC，并将其导入大肠杆菌BL21（λDE3）。用异丙基-β-D-硫代半乳糖苷（IPTG）诱导工程菌表达IpaC，通过SDS-PAGE进行鉴定，并用QIA表达纯化系统进行纯化。
SDS-PAGE显示融合蛋白条带，相对分子质量约为63 000，其表达量达到大肠杆菌BL21（λDE3）总蛋白的11%。经过适当纯化，当咪唑洗脱浓度为350 mmol/L时，目标融合蛋白的纯度达到90%以上。
重组质粒pET32a-ipaC在大肠杆菌BL21（λDE3）中已稳定高效表达，QIA表达纯化系统简单高效。
