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| 犬α-干扰素蛋白在不同表达载体中的生物学活性比较 | |
| 引用本文： | 刘欢,金红岩,侯强,史秋梅,董瑞凯,张通明,侯绍华,沈萍.犬α-干扰素蛋白在不同表达载体中的生物学活性比较[J].中国畜牧兽医,2016,43(10):2560-2565. |
| 作者姓名： | 刘欢 金红岩 侯强 史秋梅 董瑞凯 张通明 侯绍华 沈萍 |
| 作者单位： | 1. 河北科技师范学院预防兽医实验室, 秦皇岛 066003; 2. 中国农业科学院北京畜牧兽医研究所, 北京 100193 |
| 基金项目： | 公益性行业（农业）项目“宠物疫病快速诊断与疫苗研究与示范”（201303042） |
| 摘 要： | 根据GenBank中公布的犬α-干扰素（CaIFN-α）基因核酸序列，去掉信号肽后设计并合成1对引物，引入EcoRⅠ和HindⅢ酶切位点。以提取的犬肝脏DNA为模板，利用PCR技术扩增CaIFN-α基因，并分别克隆至表达载体pBV220、pET-32a（His标签）中，转化大肠杆菌BL21（DE3）原核表达系统。重组融合蛋白经SDS-PAGE及Western blotting鉴定。纯化目的蛋白，并采用MDCK/VSV微量细胞病变抑制法检测其抗病毒活性。试验结果表明，与pBV220载体连接的目的基因表达的蛋白活性较低，而与pET-32a（His标签）连接的目的基因表达的蛋白活性较高，达2.56×106 U/mL。通过分析不同载体对IFN-α的表达情况，为生产高活性的IFN奠定基础。 |
| 关 键 词： | 犬 &alpha -干扰素 原核表达 生物学活性 |
| 收稿时间： | 2016-02-22 |
The Comparison of Canine α-Interferon Antiviral Activity in Different Expression Vectors |
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| LIU Huan,JIN Hong-yan,HOU Qiang,SHI Qiu-mei,DONG Rui-kai,ZHANG Tong-ming,HOU Shao-hua,SHEN Ping.The Comparison of Canine α-Interferon Antiviral Activity in Different Expression Vectors[J].China Animal Husbandry & Veterinary Medicine,2016,43(10):2560-2565. | |
| Authors: | LIU Huan JIN Hong-yan HOU Qiang SHI Qiu-mei DONG Rui-kai ZHANG Tong-ming HOU Shao-hua SHEN Ping |
| Institution: | 1. Preventive Veterinary Laboratory of Hebei Normal University of Science and Technology, Qinhuangdao 066003, China; 2. Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beijing 100193, China |
| Abstract: | According to the sequence of canine interferon-α (CaIFN-α) gene published in GenBank,primers were designed,synthesized and introduced EcoRⅠand HindⅢ restriction sites.Using canine liver DNA as a template,CaIFN-α gene was amplified by PCR,then the gene was cloned into pBV220 and pET-32a(+) expression vector,and transformed into the BL21(DE3) expression system.Recombinant fusion protein was analysed by SDS-PAGE and Western blotting.Protein was purified and the antiviral activity was detected by MDCK/VSV cytopathic inhibition assay.The results showed that pBV220-α-IFN recombinant protein has a low activity,but the activity of pET32a-α-IFN recombinant protein was up to 2.56×106 U/mL.This study laid a foundation for production of highly active IFN. |
| Keywords: | canine interferon-α prokaryotic expression antiviral activity |
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