标题：联合G418筛选构建GFP转基因骨髓基质干细胞系的实验研究

      作者：王亮亮，孙婧，刘强和，邓铭，彭文丽    (桂林医学院附属医院耳鼻喉头颈外科，广西 桂林 541000)

      卷次： 2018年29卷20期

      【摘要】 目的 联合G418筛选构建绿色荧光蛋白(GFP)转基因SD大鼠骨髓基质干细胞(BMSCs)系，用于骨髓基质干细胞体外培养及向内耳神经细胞诱导分化的进一步研究。方法 制备大鼠骨髓基质干细胞，选用脂质体Lipofectamine TM 2000转染方法将含有pcDNA3.1-GFP的重组质粒转染至大鼠骨髓基质干细胞中，荧光显微镜检测GFP蛋白表达，通过G418筛选法建立稳定、含pcDNA3.1-GFP的单克隆骨髓基质干细胞系。结果 500 μg/mL的G418压力筛选后，获得了具有G418抗性的绿色荧光蛋白(GFP)转基因SD大鼠骨髓基质干细胞系。结论 成功地培育了G418抗性的GFP转基因大鼠骨髓基质干细胞系，为进行下一步体外特异微环境下的诱导分化提供实验基础。
      【关键词】 G418；绿色荧光蛋白；转染；骨髓基质干细胞
      【中图分类号】 R-332 【文献标识码】 A 【文章编号】 1003—6350（2018）20—2813—03

Experimental study on constructing bone marrow stromal stem cells line in transgenic SD rats expressing greenfluorescent protein by G418 screening.
WANG Liang-liang, SUN Jing, LIU Qiang-he, DENG Ming, PENG Wen-li.Otolaryngology- Head and Neck Surgery, Affiliated Hospital of Guilin Medical Unversity, Guilin 541000, Guangxi, CHINA
【Abstract】 Objective To construct bone marrow stromal stem cells (BMSCs) in transgenic SD rats ex-pressing green fluorescent protein (GFP) by G418 screening, in order to provide basis for in vitro culture of BMSCsand induced differentiation of BMSCs into inner ear nerve cells. Methods Recombinant plasmid containing pcD-NA3.1-GFP was transfected into the rats, BMSCs prepared by Lipofectamine TM 2000 transfection, and the expres-sion of GFP protein was detected by fluorescence microscopy. A line of stable and monoclonal bone BMSCs contain-ing pcDNA3.1-GFP was established by G418 screening. Results After pressure screening with 500 μg/mL G418,BMSCs with G418 resistance in transgenic SD rats expressing GFP were obtained. Conclusion The lines of BM-SCs of transgenic SD rats expressing GFP with G418 resistance were successfully cultured, which provides a basisfor the induced differentiation in vitro.
      【Key words】 G418; Green fluorescent protein (GFP); Transfection; Bone marrow stromal stem cells (BMSCs)·论 著·