Cloning of mouse albumin gene promoter and assay of its transcriptional activity in different cell lines

AIM:To study the tissue-specific function of albumin (ALB) in different cell lines. METHODS:Total DNA was extracted from mouse blood for PCR amplification of ALB promoter. The amplified fragments were inserted into the multiple cloning sites of pMD-18T vector and identified using restriction enzyme digestion and sequencing,then cloned into the expression vector pEGFP-1. pALB-EGFP and its control pEGFP-N1 was transfected into the cell lines of L02,HeLa,SW480 and Bxpc-3 by Lipofectamine 2000,respectively. The expressions of EGFP were detected using fluorescence microscopy and quantitatively with flow cytometry. RESULTS:ALB promoter was successfully cloned into the expression vector pEGFP-1. ALB promoter drove EGFP expression in transfected L02 cells and product of EGFP was observed at 72 h after transfection. However,no expression of EGFP in the cell lines of HeLa,SW480 and Bxpc3 was observed after transfection with pALB-EGFP. The control expression vector pEGFP-N1,which contains the promoter of CMV,drove the expression of EGFP very well in all these 4 cell lines after transfection. Level of EGFP transient expression driven by ALB was only a quarter of that driven by CMV promoter in L02. However,after G418 selection,the level of EGFP stable expression driven by ALB reached a considerable level as compared to that of CMV promoter. CONCLUSION:Cloned mouse ALB promoter presents a strong transcriptional activity in L02 cells among the tested cell lines. It will serve as a suitable tool for transgenic mouse in the future.