| 猪白介素4在CHO-K1细胞中的重组表达及生物学活性分析 |
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| 引用本文： | 陈国华,景志忠,房永祥,王笑笑,窦永喜,蒙学莲,宋世斌.猪白介素4在CHO-K1细胞中的重组表达及生物学活性分析[J].中国兽医学报,2008,28(12). |
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| 作者姓名： | 陈国华 景志忠 房永祥 王笑笑 窦永喜 蒙学莲 宋世斌 |
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| 作者单位： | 中国农业科学院兰州兽医研究所家畜疫病病原生物学国家重点实验室/甘肃省动物寄生虫病重点实验室,甘肃,兰州,730046 |
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| 基金项目： | 国家高技术研究发展计划(863计划)
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兰州市科技攻关项目
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| 摘 要： | 将猪白介素4(Porcine interleukin4,PIL-4)基因亚克隆到真核表达载体pEGFP-N1中,构建重组表达质粒pEGFP-PIL-4,利用脂质体法转染CHO-K1细胞,采用荧光显微镜实时观察、RT-PCR和Western-blot分别检测CHO细胞转录表达目的分子的情况,通过MTT法检测所表达蛋白的生物学活性。结果在将重组质粒转染CHO-K1细胞中24、48 h后的均观察到绿色荧光;经G418筛选14~20 d后转染的细胞形成了阳性细胞集落,用RT-PCR扩增出约339 bp的目的基因片段;经Western-blot检测到约为39 000的特异蛋白分子条带,并证明在转染重组质粒的细胞中表达了高生物活性的重组蛋白。
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| 关 键 词： | 白介素4 真核表达 CHO细胞 生物活性 |
Expression and bioactive analysis of porcine interleukin 4 in the CHO-KI cells |
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| CHEN Guo-hua,JING Zhi-zhong,FANG Yong-xiang,WANG Xiao-xiao,DOU Yong-xi,MENG Xue-lian,SONG Shi-bin.Expression and bioactive analysis of porcine interleukin 4 in the CHO-KI cells[J].Chinese Journal of Veterinary Science,2008,28(12). |
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| Authors: | CHEN Guo-hua JING Zhi-zhong FANG Yong-xiang WANG Xiao-xiao DOU Yong-xi MENG Xue-lian SONG Shi-bin |
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| Abstract: | The IL-4 gene of porcine was subcloned into the eukaryotic expression vector pEGFP-N1 to construct the recombinant plasmid pEGFP-PIL-4,which was transfected into the CHO-K1 cells by the method of using Lipofectin,The expression of PIL-4 was determined by fluoroscopy,RT-PCR and Western-blot.After 24 hours and 48 hours,the Green fluorescence can be observed under fluorescence microscope and a 339 bp cDNA was detected by RT-PCR after screening 14 days with G418,39 000 protein was detected by Western-blot.The results indicate that fusion protein of PoIL-4 and EGFP was successfully expressed in CHO-K1 cells.And expressed protein had obviously biological activity. |
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| Keywords: | interleukin4 eukaryotic expression CHO cell bioactivity |
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