医学分子生物学杂志2011，Vol.8Issue(1)：1-6,6.DOI:10.3870/j.issn.1672-8009.2011.01.001
糖基因Glt8d2重组蛋白的表达、纯化及多克隆抗体的制备
Expression,Purification and Polyclonal Antibody Preparation for A Novel Glycogene Glt8d2
摘要
Abstract
Objective To express and purify glycogene Glt8d2 recombinant protein. and to prepare rabbit polyclonal antibody specific for Clt8d2 recombinant protein. Methods Glt8d2 cDNA was ligated into the prokaryotic expressive vector pET-32a ( + ) . The resultant plasmid was transformed into E. coil BL21 ( DE3 ) . Glt8d2 recombinant protein expression was induced under IPTG condition and analyzed with western blotting, followed by Ni + affinity column purification. The Glt8d2 recombinant protein product was further confirmed by mass spectrometry. New Zealand rabbits were immunized with the purified pET-32a ( + ) -Glt8d2 fusion protein to gain polyclonal antibody. Antibody specificity and potency were evaluated by Western blot and ELISA. Results The Glt8d2 fusion protein was highly expressed. The protein was produced mainly in the inclusion body. ELISA showed the titer of the polyclonal antibody higher than 1∶320000. Western blot confirmed the high specificity of Glt8d2 antibodies. Immunohistochemistry staining of liver tissue showed that this glycosyltransferase are mainly scattered in the cytoplasm of hepatocytes. Conclusion The recombinant Glt8d2 fusion protein and the Glt8d2 specific polyclonal antibody will be useful for investigation of Glt8d2 biological function.关键词
糖基因/糖基转移酶/脂肪肝/载脂蛋白/糖基化修饰分类
生物科学引用本文复制引用
魏洪莲,张锦前,肖凡,李学永,成军,魏红山,展玉涛..糖基因Glt8d2重组蛋白的表达、纯化及多克隆抗体的制备[J].医学分子生物学杂志,2011,8(1):1-6,6.基金项目
国家自然科学基金(No.81041017,30872243) （No.81041017,30872243）
