| Ataxin-3融合蛋白表达、纯化及抗血清的制备 |
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| 引用本文： | 肖涵,胡治平,汤建光,谭洁琼.Ataxin-3融合蛋白表达、纯化及抗血清的制备[J].现代生物医学进展,2009,9(14). |
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| 作者姓名： | 肖涵 胡治平 汤建光 谭洁琼 |
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| 作者单位： | 1. 中南大学湘雅二医院神经内科,湖南,长沙,410011
2. 中南大学医学遗传学国家重点实验室,湖南,长沙,410078 |
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| 基金项目： | 国家自然科学基金资助(30670720) |
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| 摘 要： | 目的:表达GST-ataxin-3-N融合蛋白并制备GST-ataxin-3特异性抗体,为深入研究其功能及其在SCA3发病机制中的作用提供重要的技术和材料保障.方法:将人ataxin-3氨基端基因克隆入原核表达载体pGEX-4T-2,在大肠杆菌(E.coli)BL21中表达,用Glutathione sepharose4B凝胶亲和柱纯化目的蛋白.利用纯化的GST-ataxin-3-N蛋白制备多克隆抗体.结果:成功构建了原核表达载体,得到高表达量的融合蛋白,经亲和层析柱纯化获得较高纯度的GST-ataxin-3-N融合蛋白.以融合蛋白免疫新西兰兔得到Ataxin-3-N多克隆抗体,Western Blotting及免疫荧光均证实该抗体能够识别Ataxin-3-myc蛋白,具有较高特异性.结论:利用原核表达人GST-ataxin-3-N融合蛋白制备的Ataxin-3多克隆抗体具有较好的特异性,可用于该蛋白的相关研究.
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| 关 键 词： | ataxin-3 融合蛋白 多克隆抗体 |
Expression and purification of Ataxin-3 fusion protein in E.coli and preparation of polyclonal antibody against ataxin-3 |
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| XIAO Han,HU Zhi-ping,TANG Jian-guang,TAN Jie-qiong.Expression and purification of Ataxin-3 fusion protein in E.coli and preparation of polyclonal antibody against ataxin-3[J].Progress in Modern Biomedicine,2009,9(14). |
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| Authors: | XIAO Han HU Zhi-ping TANG Jian-guang TAN Jie-qiong |
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| Institution: | XIAO Han1,HU Zhi-ping1,TANG Jian-guang1,TAN Jie-qiong2 (1 Department of Neurology,second Xiangya Hospital,Central South University,changsha 410011 China 2 National Key Laboratory o f Medical Genetics,changsha 410078,China) |
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| Abstract: | Objective: To obtain ataxin-3 fusion protein and prepare anti-ataxin-3 polyclonal antibody. Methods: The DNA fragments encoding N-terminal 177 amino acids of ataxin-3 was amplified by PCR and cloned into a prokaryotic expression vector, pGEX-4T-2 , and transformed into E.coli BL21. The expressed proteins were purified from total proteins with Glutathione sepharose 4B agarose column. The New Zealand rabbits were immunized with the purified fusion proteins to prepare polyclonal antiserum. Results: Prokaryotic... |
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| Keywords: | Ataxin-3 Fusion protein Polyclonal antibody |
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