摘要
目的建立以增强型绿色荧光蛋白基因示踪骨髓间质干细胞的方法.方法采用密度梯度离心法分离、培养兔骨髓间质干细胞,以脂质体介导法转染增强型绿色荧光蛋白基因的表达质粒,荧光显微镜观察基因表达及转染效率.结果绿色荧光蛋白在基因转染12 h后开始表达,48～72 h达高峰,并在1周内有较强的表达,以后逐渐减弱,4周左右仍有少量表达,转染效率与质粒、脂质体的浓度有关,外缘基因导入后不影响骨髓间质干细胞的生长和增殖.结论脂质体介导的绿色荧光蛋白基因转染骨髓间质干细胞后能安全、有效地表达,转染效率为20%～30%,是一种较为理想的干细胞示踪方法.
Aim To establish a method for tracking bone marrow mescnchymal stem cells （MSC） through transfection of enhanced green fluorescence protein （EGFP） gene, Methods Mesenehymal stem cells were isolated from bone marrow and purified by density centrifuge in vitro, The cultured cells were transferred with the expressing plamid of EGFP （pEGFP） by means of lipofectamine media methods. The transient expression and transfection efficiency were subsequently observed by fluorescent microscopy, Results The expression of EGFP was initially found in 12 h after transfection, reached optimal state in 48-72 h, remained strong express within 1 week and than becarne weak for 4 weeks, The efficiency of transfection was corrclated with the concentration of plamid and lipofectamine respectively. Particularly, the growth and proliferation of rnesenchymal stem cells wasn＇t disturbed by EGFP gene transfection. Conclusions EGFP transfected by lipofectamine media could be safely expressed in MSC in vitro. The efficiency of transfection could reach 20% ～ 30% with proper concentration of plamid and lipofactamine. EGFP as a report gene to label stem cells was an ideal method for tracking.
出处
《中国动脉硬化杂志》
CAS
CSCD
2005年第5期549-552,共4页
Chinese Journal of Arteriosclerosis
基金
西安交通大学教育振兴行动计划(XY10082008)
