| 猪氨基肽酶在昆虫细胞的分段表达及鉴定 |
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| 引用本文： | 单智夫,唐丽杰,乔薪媛,葛俊伟,李一经*.猪氨基肽酶在昆虫细胞的分段表达及鉴定[J].东北农业大学学报,2012(9):12-16. |
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| 作者姓名： | 单智夫 唐丽杰 乔薪媛 葛俊伟 李一经* |
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| 作者单位： | 东北农业大学动物医学学院,哈尔滨 150030 |
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| 基金项目： | 国家自然科学基金(30671574) |
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| 摘 要： | 为研究猪的氨基肽酶(Porcine aminopeptidase, pAPN)的特异性功能区域及作用机制,将除去信号肽的pAPN分为SPA,SPB和SPC三段,利用一对同尾酶Xhol I和Sal I将信号肽分别与SPA,SPB和SPC相连接后再构建到杆状病毒表达载体pFastBacTM1上,通过杆状病毒-昆虫细胞表达体系进行蛋白表达.结果表明,经SDS-PAGE分析可见获得的目的蛋白与预期大小相一致,即SPA约为42 ku,SPB和SPC相同,约为56 ku的分泌蛋白;经western-blot鉴定三段蛋白均可与天然的pAPN抗血清特异性结合,表明已经成功获得SPA,SPB和SPC三段蛋白,可为进一步研究pAPN特异性功能区域奠定基础.
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| 关 键 词： | pAPN分段 杆状病毒-昆虫细胞表达体系 表达与鉴定 |
Expression and identification of the subsection from pAPN in insect cells expression systems |
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| SHAN Zhifu, TANG Lijie, QIAO Xinyuan, GE Junwei, LI Yijing.Expression and identification of the subsection from pAPN in insect cells expression systems[J].Journal of Northeast Agricultural University,2012(9):12-16. |
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| Authors: | SHAN Zhifu TANG Lijie QIAO Xinyuan GE Junwei LI Yijing |
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| Institution: | (School of Veterinary Medicine,Northeast Agricultural University,Harbin 150030,China) |
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| Abstract: | To investigate which part of porcine aminopeptidase(pAPN) is the functional domain of receptor for PEDV,we removed the signal peptide and divided pAPN into three sections: SPA,SPB and SPC.Then we use a pair of isocaudarner Xhol I and Sal I to connect the signal peptide and SPA,SPB and SPC respectively and connected to pFastBacTM1 finally.We express the SPA,SPB and SPC by Bac-to-Bac baculovirus expression systems and get the secreted protein 42,56 ku as we expected by SDS-PAGE.The result of western blot shows that SPA,SPB and SPC can specific binding to the nature antiserum of pAPN.It shows that we have get the three parts of pAPN successfully and prepared for the futher research on the functional domain of pAPN. |
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| Keywords: | subsection of pAPN insect cells expression systems expression and identifica-tion |
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