摘要
目的构建带加强型绿色荧光蛋白 (EGFP)基因的人骨形成蛋白 2 (hBMP2 )真核表达载体 ,观察其在正常人皮肤成纤维细胞中的表达情况。方法构建携带hBMP2和EGFP基因的真核表达载体 pcDNA3 hBMP2 IRES EGFP(pcBE) ,将其转染NIH3T3细胞和正常人皮肤成纤维细胞后 ,采用RT PCR和免疫组化染色方法分别检测人BMP2和EGFP基因在mRNA和蛋白质水平上的表达情况。结果转染后的NIH3T3和正常人皮肤成纤维细胞均能转录人BMP2mRNA和表达人BMP2蛋白 ,同时显示绿色荧光。结论 pcBE真核表达载体可在正常人皮肤成纤维细胞内有效表达人BMP2蛋白 ,并以其荧光报告基因起示踪作用。
Objective To construct an eukaryotic expressing vector for recombinant human bone morphogenetic protein2 (rhBMP2) that contains enhanced green fluorescence protein (EGFP), and to determine its expression in the human skin fibroblasts. Methods The rhBMP2 eukaryotic expressing vector pcDNA3-hBMP2-IRES-EGFP (pcBE) was constructed and transfected to NIH3T3 and normal human skin fibroblasts. The transient transfection and expression of pcBE in the two types of cells were determined by RT-PCR and immunohistochemical methods. Results Expression of both hBMP2 and EGFP in mRNA and protein levels could be detected, and green fluorescence could be visualized in the transfected cells. Conclusion NIH3T3 and normal human skin fibroblasts, showing green fluorescence, could effectively transcript BMP2 mRNA and express BMP2 protein. This study provides feasibility for tracing the transplanted fibroblasts expressing BMP2 in a bone healing fracture model.
出处
《上海第二医科大学学报》
CSCD
2004年第12期1007-1010,共4页
Acta Universitatis Medicinalis Secondae Shanghai
基金
国家自然科学基金 (30 1 70 943)资助项目
