| 人白细胞介素4的原核表达、复性、纯化及生物学活性鉴定 |
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| 引用本文： | 李炯,崔凯军,文静,赵志伟,陈平,田聆,阚兵,文艳君,邓洪新,范琳玉,魏于全. 人白细胞介素4的原核表达、复性、纯化及生物学活性鉴定[J]. 生物医学工程学杂志, 2007, 24(4): 866-869 |
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| 作者姓名： | 李炯 崔凯军 文静 赵志伟 陈平 田聆 阚兵 文艳君 邓洪新 范琳玉 魏于全 |
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| 作者单位： | 1. 四川大学,华西医院,生物治疗国家重点实验室,成都,610041;四川大学,生命科学学院,成都,610064
2. 四川大学,华西医院,生物治疗国家重点实验室,成都,610041 |
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| 摘 要： | 用PCR方法从pORF-hIL4质粒中扩增重组人白细胞介素4(hIL-4)的基因,构建PQE60原核表达载体并诱导其目的蛋白的表达。对表达的目的蛋白的可溶性进行鉴定,发现hIL-4基因表达产物在大肠杆菌中主要以不溶性包涵体形式存在,经过超声波破细菌、包涵体提取、洗涤处理后,用5mol/L盐酸胍变性溶解包涵体沉淀,上清稀释复性后透析,再经镍亲和层析进行纯化。纯化后的hIL-4进行TF-1细胞体外增生实验检测其生物学活性后,可用于人外周血树突状细胞的体外诱导培养。
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| 关 键 词： | 重组人白细胞介素4 包涵体 复性 纯化 生物学活性 树突状细胞 |
| 修稿时间： | 2005-04-112005-06-27 |
Prokaryotic Expression, Purification, Refolding and Biological Assays of Recombinant Human Interleukin 4 |
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| Li Jiong,Cui Kaijun,Wen Jing,Zhao Zhiwei,Chen Ping,Tian Ling,Kan Bing,Wen Yanjun,Deng Hongxin,Fan Linyu,Wei Yuquan. Prokaryotic Expression, Purification, Refolding and Biological Assays of Recombinant Human Interleukin 4[J]. Journal of biomedical engineering, 2007, 24(4): 866-869 |
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| Authors: | Li Jiong Cui Kaijun Wen Jing Zhao Zhiwei Chen Ping Tian Ling Kan Bing Wen Yanjun Deng Hongxin Fan Linyu Wei Yuquan |
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| Affiliation: | 1.State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu 610041, China;2.School of Life Science, Sichuan University, Chengdu 610064, China |
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| Abstract: | A DNA fragment encoding human interleukin 4 was obtained by PCR from pORF-hIL4 plasmid. The amplified fragment was inserted into prokaryotic expression vector PQE60 and recombinant protein was expressed in E. Coli M15 by adding isopropyl-beta-D-thiogalactoside (IPTG). The hIL-4 protein was present as insoluble inclusion bodies in the bacterial extract. After denaturation of inclusion bodies with 5 mol/L guanidine hydrochloride, the supernate was diluted to get renaturized. Then dialysis and Ni chelating chromatography were used for purification. TF-1 proliferation assay of recombinant human interleukin 4 was performed, and then rhIL-4 was fit to be used for proliferation of human dendritic cells from monocyte in vitro. |
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| Keywords: | Recombinant human interleukin 4(rhIL-4) Inclusion body Refolding Purification Biological activity Dendritic cells |
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