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目的 研究IL－18在大肠杆菌中高效表达及纯化工艺，并诱导KG－1 细胞产生IFN－γ。 方法利用RT－nPCR从正常人胎肝组织中扩增获得人IL－18基因，克隆于pThiohisA载体，转化宿主菌BL－21，1mmol/LIPTG；诱导，表达产物以包涵体形式存在，经2％Triton X－100洗涤，在 8mol/L尿素下变性阴离子柱层析，透析复性后再经凝胶过滤纯化，通过纯化产物诱导入骨髓瘤单核细胞 KG－l（ATCC CCL－246）产生 IFN－γ，并检测其生物学活性。结果 获得了在大肠杆菌中的稳定高效表达，表达的目的蛋白占菌体总蛋白的25％左右，纯化后纯度可达95％以上，具有显著的刺激细胞产生IFN－γ的活性。结论 制备具有生物学活性高纯度的rhIL－18方法的建立，为基础研究与临床应用开发奠定了基础。
Objective To study the efficient expression and purification of IL-18 in Escherichia coli and the induction of IFN-γ production by KG-1 cells. Methods The human IL-18 gene was amplified from normal human fetal liver tissue by RT-nPCR and cloned into pThiohisA vector. The recombinant plasmid was transformed into BL21 and lmmol / L IPTG. The expressed product was expressed in inclusion bodies. After 2% Triton X-100, denatured by 8 mol / L urea, and purified by gel filtration after dialysis refolding. The purified product was induced into myeloma monocytic KG-1 (ATCC CCL-246) -γ, and tested for biological activity. The results showed that the target protein expressed in Escherichia coli was about 25% of the total bacterial protein, and the purity of purified protein was over 95% after purification. It has the remarkable activity of stimulating IFN-γ production by cells. Conclusion The preparation of rhIL-18 with high biological activity and purity has laid a foundation for the development of basic research and clinical application.