人角质细胞生长因子2在不同原核表达系统中的表达差异

目的:比较重组人角质细胞生长因子2(rhKGF-2)在不同原核表达系统中的表达量和表达稳定性之间的差异。方法:利用PCR方法从人胚胎肺成纤维细胞cDNA扩增得到hKGF-2序列,双酶切后分别克隆到pBV220、pQE31和pET-24b载体中,分别转化大肠杆菌JM109、M15和BL21(DE3)进行诱导表达,SDS-PAGE分析rhKGF-2的表达量和表达稳定性,并纯化rhKGF-2。结果:pBV220-rhKGF-2与pET-24b-rhKGF-2在宿主菌内经诱导后均有目的蛋白表达,其中pBV220-rhKGF-2的表达量约占菌体总蛋白的10%,pET-24b-rhKGF-2的表达量约占菌体总蛋白的25%,且均为可溶性表达,但后者的表达稳定性明显优于前者,而pQE31-rhKGF-2在宿主菌内几乎没有表达。结论:hKGF-2在不同原核表达系统中的表达量和表达稳定性存在明显差异。

Comparison of hKGF-2 Expression in Different Prokaryotic Expression Systems

Objective： To compare expression level and stability of human keratinocyte growth factor 2（hKGF-2） in different prokaryotic systems. Methods： The hKGF-2 gene was amplified by PCR from cDNA of human embryonic lung fibroblast. After enzyme digestion, the hKGF-2 gene was cloned into plasmid pBV220, pQE3t and pET- 24b and then transformed into different host bacteria E.coli JM109, M15 and BL21（DE3） respectively. The hKGF- 2 quantity and stability was determined by SDS-PAGE. Results： The expression level of hKGF-2 in pBV220 was up to 15% of total bacterial proteins. The expression level of protein of interest in pET-24b transformed E.coli BL21 （DE3） reached 25%. And both of them were soluble, however, the former was more stable. But there was no observed expression of pQE31-rhKGF-2 in E.coli M15. Conclusion： The expression level and expression stabili- ty of hKGF-2 displayed significant difference in different prokaryotic expression systems.