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GAS41蛋白的表达、抗体制备及其亚细胞定位
激光生物学报摘要 2005-6
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丁小凤 , 胡 翔, 张 健*
(湖南师范大学生命科学学院，中国湖南 长沙 410081)
摘 要:GAS41是新发现的与神经胶质瘤密切相关的基因。为了对该基因进行深入的功能研究，成功地将GAS41基因构建于原核表达载体pQE-N3,转化BL21（DE3）获得融合表达产物。对含融合蛋白的包含体进行溶解和复性,通过免疫新西兰大白兔获得兔抗GAS41多克隆抗体。采用Western印迹技术,用该抗体检测GAS41基因的原核和真核表达产物,证明该抗体有较好的针对GAS41蛋白的专一性,可用于对GAS41的结构和功能研究。同时,用该抗体通过荧光免疫细胞进行定位分析发现,GAS41蛋白均匀分布于COS7细胞的细胞核中,提示GAS41可能是一个重要的转录因子。
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Subcellular Location of GAS41 by its Expression and Preparation of Antiserum
DING Xiao-feng, HU Xiang, ZHANG Jian*
（College of Life Science, Hunan Normal University， Changsha 410081, Hunan, China）
Abstract:GAS41 is a recently identified gene slightly related to glioma. In order to study the expression, subcellular distribution and the function of GAS41 protein，the coding region of human GAS41 gene was successfully subcloned into the prokaryotic expression vector pQE-N3，The vector was then introduced into E. coli BL21（DE3）cells for highly efficient expression. His-fusion protein of 30 kD was detected in the inclusion body. After refolded and purified，this protein was used to immunize new Zealand white rabbit to produce polyclonal antibodies. The specificity of the anti-GAS41 antibody was successfully proved by Western blotting and analysis on both of the prokaryotic and eukaryotic expressed products of GAS41 protein.Moreover, the GAS41 immunoreactive cells were found to be uniformly distributed in the nucleolus,this suggests GAS41 may be an important transcription factor.
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Bioinformatics,
sequence analysis; GCG; Life Science News; Drug Discovery.
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