| 猪流行性腹泻病毒M蛋白的原核表达 |
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| 引用本文： | 刘梦茜,李春燕,陈仕怡,袁晓琴,星东,王全溪.猪流行性腹泻病毒M蛋白的原核表达[J].福建农林大学学报(自然科学版),2017,46(3). |
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| 作者姓名： | 刘梦茜 李春燕 陈仕怡 袁晓琴 星东 王全溪 |
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| 作者单位： | 福建农林大学动物科学学院,福建 福州,350002 |
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| 基金项目： | 福建农林大学科学发展基金资助项目 |
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| 摘 要： | 本试验设计了M基因的特异性引物,扩增了M基因,成功构建了重组质粒,并转化至Escherichia coli BL21(DE3)感受态细胞中,优化了异丙基-β-D-硫代半乳糖苷(IPTG)诱导表达条件.结果表明,重组表达的融合蛋白Pet-32a-M大小约为43 ku,与预期大小相符.十二烷基硫酸钠聚丙烯酰胺凝胶电泳(SDS-PAGE)检测表明,M蛋白为包涵体蛋白,且在IPTG浓度为0.8 mmol·L-1,温度为37℃,诱导时间为8 h的条件下,蛋白表达效果最佳.蛋白免疫印迹检测结果进一步显示,重组蛋白Pet-32a-M在体外可以被成功表达.
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| 关 键 词： | 猪流行性腹泻病毒 M蛋白 原核表达 |
Prokaryotic expression of porcine epidemic diarrhea virus M protein |
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| LIU Mengxi,LI Chunyan,CHEN Shiyi,YUAN Xiaoqin,XING Dong,WANG Quanxi.Prokaryotic expression of porcine epidemic diarrhea virus M protein[J].Journal of Fujian Agricultural and Forestry University,2017,46(3). |
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| Authors: | LIU Mengxi LI Chunyan CHEN Shiyi YUAN Xiaoqin XING Dong WANG Quanxi |
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| Abstract: | To increase the expression level of M protein which is a candidate antigen for porcine epidemic diarrhea virus, specific primer for M gene was designed and amplified. The positive recombinant plasmid was identified and transformed into Escherichia coli BL21 ( DE3) competent cell, and followed by being induced by isopropylβ-D-thiogalactoside ( IPTG) . Then the inducible expres-sion system was optimized in terms of IPTG concentration, expression time and temperature. Results showed that the recombinant protein was successfully expressed, with the molecular weight being about 43 ku. SDS-Polyacrylamide gelelectrophoresis ( SDS-PAGE) analysis indicated that M protein is an inclusion body protein. The optimal induction conditions were to add 0.8 mmol·L-1 IPTG and be induced at 37 ℃ for 6 h. In addition, Western-blot contfirmed that the recombinant plasmid Pet-32a-M was trans-formed into E.coli BL21 (DE3) competent cell and successfully expressed in vitro. |
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| Keywords: | porcine epidemic diarrhea virus membrane protein procaryotic expression |
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