Isolation and identification of bone marrow mesenchymal stem cells from transgenic rats with green fluorescent protein gene

Wang Yang; Cao Zhi-qiang

doi:10.3969/j.issn.2095-4344.2014.37.004

Abstract：BACKGROUND:Mesenchymal stem cells from transgenic animals carry green fluorescent protein that can be stably expressed in viable cells and can be used to rapidly screen modified cells in comparison with traditional virus and plasmids. OBJECTIVE:To study the biological features of bone marrow derived stromal cells from transgenic rats with green fluorescent protein gene. METHODS:Bone marrows were isolated from the bilateral long bones of 2-week-old transgenic rats with green fluorescent protein gene, and cultivated in conditioned culture medium to obtain bone marrow mesenchymal stem cells with green fluorescent protein gene. The passage 5 bone marrow mesenchymal stem cells with green fluorescent protein gene were analyzed by flow cytometry to detect cellsurface molecules, and were induced in calcium or adipogenic induction medium. After calcium induction, the alizarin red staining was performed to test the formation of calcium concentration. After adipogenic induction, bone marrow mesenchymal stem cells were detected with oil red O staining. RESULTS AND CONCLUSION:Bone marrow mesenchymal stem cells, stably expressing green fluorescent protein gene, were obtained successful y from the transgenic rat with green fluorescent protein gene. The cells expressed CD90, CD105 strongly, but did not express or weakly expressed CD14 andCD45. After 3 weeks of calcium induction, jacinth calcium salts could be detected in the cells by alizarin red staining. At 3 weeks of adipogenic induction, the cells showed positive oil red O staining. The green fluorescent protein gene, as report gene, exerts no effect on the cellsurface molecules and multilineage potential of bone marrow mesenchymal stem cells, which can be used as an efficient tracer.