解放军医学杂志2011，Vol.36Issue(1)：35-38,4.
大鼠脑胶质瘤荧光素酶动物模型建模细胞株C6-Luc的构建
Construction of rat glioma cell line C6-Luc for reproducing an animal model with stable expression of luciferase
摘要
Abstract
Objective To construct the rat glioma cell line C6-Luc to stably express the firefly luciferase.Methods The optimal concentration of hygromycin for screening C6 rat glioma cells was determined by concentration gradient method.The eukaryotic plasmid pGL4.50 expressing luciferase was transfected into C6 cells by using FuGENE* HD transfection reagent, followed by screening the polyclonal cell lines with hygromycin, subsequently screening the monoclonal cell line by limited dilution.The positive monoclonal cell lines were identified with reporter gene assay, thereafter the expression stability of luciferase was investigated in the positive cell lines.The bioluminescence detection in vitro in the positive monoclonal cell line was performed to determine the minimum detection amount of cells,and the correlation between bioluminescence intensity and ce11 amount was analyzed by linear regression analysis.The positive monoclonal cells were implanted into the brain of Wistar rats, and the tumor growth in rats brain was detected in vivo using the bioluminescence imaging detection system.Results The optimal concentration of hygromycin used in screening C6 cells was 250 μg/mL The eukaryotic plasmids pGL4.50 was successfully transfected into C6 cells, and 12 monoclonal cell lines were obtained by anti-hygromycin screening.A positive clone with the highest activity of luciferase, designated as C6-Luc, was successfully identified by using luciferase reporter gene assay, which showed a stable activity of expressing luciferase after 3 continuous passages of cultivation.The bioluminescence detection in vitro showed that the minimum detection amount of C6-Luc cells was 78.A good linear correlation existed between bioluminescence intensity and the amount of C6-Luc cells, with an equation of y=81.348x-2143.1 and correlation coefficient(r) of 0.997.The in vivo bioluminescence imaging detection showed tumorigenesis could be detected after implantation of C6-Luc cells into the brain of Wistar rats.The intensity of bioluminescence increased gradually with volume augmentation of the tumor in rats' brains.Conclusion The glioma cell line C6-Luc of rats expressing stably the firefly luciferase has been successfully constructed.关键词
荧光素酶类,萤火虫/生物发光成像/神经胶质瘤/模型,动物Key words
luciferases/ firefly/ bioluminescence imaging/ glioma/ models/ animal分类
医药卫生引用本文复制引用
黄伟,吕明,李保卫,王玉丽,邵荣光,高钟镐..大鼠脑胶质瘤荧光素酶动物模型建模细胞株C6-Luc的构建[J].解放军医学杂志,2011,36(1):35-38,4.基金项目
国家自然科学基金(30873168) （30873168）
