| 大鼠ANT1蛋白的原核表达纯化及多克隆抗体的制备 |
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| 引用本文： | 孟喜君,李传坤,王茂德,谢万福.大鼠ANT1蛋白的原核表达纯化及多克隆抗体的制备[J].第四军医大学学报,2009(21):2275-2277. |
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| 作者姓名： | 孟喜君 李传坤 王茂德 谢万福 |
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| 作者单位： | 西安交通大学医学院第一附属医院神经外科; |
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| 摘 要： | 目的：表达和纯化大鼠腺苷酸转运体（ANT1）蛋白并制备其多克隆抗体.方法：通过PCR扩增大鼠脑组织中ANT1的编码区序列,构建ANT基因重组载pET28a-ANT1,转化至E. coli BL21（DE3）,IPTG诱导表达,Ni2＋-NTA离子交换树脂层析纯化;纯化的ANT1蛋白免疫新西兰大白兔制备多克隆抗体并亲和纯化,并通过Western Blot法鉴定其特异性.结果：成功构建重组质粒pET28a-ANT1,经IPTG诱导在大肠杆菌中表达了Mr约为32×10^3的目的蛋白;亲和层析纯化后免疫的新西兰大白兔获得其多克隆抗体;曝光后胶片上均可见Mr32×10^3的阳性条带,与纯化的大鼠ANT1重组蛋白处于同一水平,表明纯化的抗ANT多克隆抗体具有高度的特异性.结论：利用基因重组技术,在大肠杆菌中成功表达了大鼠ANT1融合蛋白并制备了其多克隆抗体,为研究腺苷酸转运体蛋白在癫痫发病过程中的作用机制奠定了基础.
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| 关 键 词： | 原核表达 多克隆抗体 腺苷酸转运体1 |
Expression,purification of rat ANT1 protein and preparation of its polyclonal antibody |
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| MENG Xi-Jun,LI Chuan-Kun,WANG Mao-De,XIE Wan-Fu.Expression,purification of rat ANT1 protein and preparation of its polyclonal antibody[J].Journal of the Fourth Military Medical University,2009(21):2275-2277. |
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| Authors: | MENG Xi-Jun LI Chuan-Kun WANG Mao-De XIE Wan-Fu |
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| Institution: | MENG Xi-Jun,LI Chuan-Kun,WANG Mao-De,XIE Wan-Fu Department of Neurosurgery,First Affiliated Hospital,Medical School,Xi'an Jiaotong University,Xi'an 710061,China |
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| Abstract: | AIM:To express and purify the adenine nucleotide transporte protein (ANT1)and to prepare its polyclonal antibody. METHODS:The CDS of ANT1 was amplified by PCR from rat,and then subcloned into prokaryotic expression vector pET28a ,after transformed with the vector,the BL21(DE3)stains were induced by IPTG. The expression product was purified by Ni2+-NTA ion exchange resin. The purified protein was analyzed by SDS-PAGE,and then immuned the New Zealand white rabbits to prepare antibody. The polyclonal antibody ... |
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| Keywords: | prokaryotic expression polyclonal antibody adenine nucleotide transporte 1 |
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