Establishment of an Osteoblastic Model Reflecting Cbfal Activity with Fluorescent Intensity

Abstract：Objective To obtain MG63 cell lines which are stably transfected with enhanced green fluorescent protein(EGFP) reporter gene drived by 6OSE2 promoter and select the cell strain which can reflect Cbfal activity by EGFP fluorescence. Methods 6OSE2 promoter was synthesized and cloned into pUC57-T vector. The 140 bp promoter fragment inserted into deleted CMV-promoter pEGFP-Nl vector to construct eukaryotic expression vector. MG63 cell was transfected with this vector by Lipofectamine 2000 and stably selected by G418. Analysed the EGFP fluorescence intensity of the stably transfected MG63 cell stain and detection of ALP activity after treatment with different concentration of IGF-I or VD_3 ,then selected one which could reflecting Cbfal activity. Results The pUC-6OSE2 vector and p6OSE2-EGFP expression vector were constructed. Through stably transfection and selection, one OSE-MG63 cell strain was gained,which fluorescence intensity could reflect the treatment of IGF-I or VD_3. Conclusion The osteoblastic model which can reflect Cbfal activity is established successfully. This cell line will be useful for studying the effects of microgravity on the activity of Cbfal and the signal pathways related with bone form.