【摘要】 目的:构建再生基因4(Reg Ⅳ)的原核表达载体,获得重组人Reg Ⅳ蛋白。方法:将已构建好的pEGFP-C1-Reg Ⅳ质粒进行酶切纯化获得目的基因,并构建到pET-30a原核表达载体上,经测序鉴定后将序列正确的重组质粒转化E.coli BL21(DE3),IPTG诱导表达,SDS-PAGE凝胶电泳鉴定,并用凝胶层析对表达产物进行复性纯化。结果:构建了pET-30a-Reg Ⅳ原核表达重组质粒,IPTG诱导表达出约22KD的重组蛋白,表达产物经复性纯化后获得纯度较高的目的蛋白。结论:成功构建了Reg Ⅳ原核表达载体,并成功表达了重组人Reg Ⅳ蛋白。更多还原

【Abstract】 Objective:To construct the prokaryotic expression system of regenerating gene Ⅳ(Reg Ⅳ)and obtain recombinant human Reg Ⅳ protein. Methods:The constructed pEGFP-C1-RegⅣ plasmid is digested and purified to obtain the target gene,and constructed into the pET-30a prokaryotic expression vector. After sequencing and identification,the recombinant plasmid with the correct sequence was transformed into E.coli BL21(DE3),and was induced by isopropyl-beta-D-thiogalactoside(IPTG)to expression. The expressed product was identified by SDS-PAGE gel electrophoresis,and was renatured and purified by gel chromatography. Results:The recombinant expression vector pET-30a-RegⅣ was constructed and about 22 KD recombinant protein was induced. The target protein with higher purity was obtained after renaturation and purification. Conclusion:The prokaryotic expression vector pET-30a-RegⅣ was successfully constructed,and the recombinant human Reg Ⅳ protein was successfully expressed in prokaryotic expression system.更多还原