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亚洲牛带绦虫泛素缀合酶E2的克隆表达及免疫鉴定
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亚洲牛带绦虫泛素缀合酶E2的克隆表达及免疫鉴定
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Journal Article
亚洲牛带绦虫泛素缀合酶E2的克隆表达及免疫鉴定
2009
Overview
目的 克隆表达亚洲牛带绦虫泛素缀合酶E2（UBCE2）全长编码序列，获得纯化的重组蛋白，为进一步功能研究做充分准备。方法以亚洲牛带绦虫成虫cDNA文库中含有UBCE2基因的质粒作为模板，扩增该基因，将其克隆到原核表达载体pET-28a（＋）中，测序鉴定重组质粒，在大肠杆菌BL-21／DE3中用IPTG诱导，表达产物通过十二烷基磺酸钠一聚丙烯酰胺凝胶电泳（SDS-PAGE）鉴定，重组后的蛋白用His-镍蛋白纯化柱纯化，并使用Western blotting分析其免疫反应性。结果PCR、双酶切及DNA测序结果均表明重组质粒pET-28a（＋）-UBCE2构建成功，该基因在大肠杆菌中以包涵体形式得到表达，十二烷基肌氨酸钠纯化后蛋白通过SDS-PAGE鉴定结果表明该基因在大肠杆菌BL-21／DE3得到高效表达，亲和层析得到高纯度的蛋白且该重组蛋白可被亚洲带绦虫及牛带绦虫患者血清识别，表明其有免疫反应性。培论亚洲牛带绦虫UBCE2可在原核表达系统中表达，并具有免疫反应性，为进一步研究该基因的功能奠定基础。
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