关键词: CD133启动子, 加强型绿色荧光蛋白, 肿瘤干细胞, Hep-2细胞系,  

Abstract: Objective To construct the  gene expression vector  of  enhanced green fluorescent protein （eGFP）regulated by human CD133 gene promoter and to observe the expression ability of the human CD133 gene promoter in laryngocarcinoma Hep-2 cell line,and to provide experimental basis for the target therapy of tumor stem cells(CSCs) regulated by  human CD133 gene promoter.  Methods The CD133 gene promoter was cloned from the genomic DNAs of human peripheral blood by PCR method and  cloned into lentivirus pWPXLd-eGFP by using gene recombination technology to construct eGFP gene expression vector induced by human CD133 gene promoter. The recombined vector was identified by PCR and enzyme digestion. The recombined vector was transfected into Hep-2 cells induced by liposome. The cells were divided into  blank control group (non-transfected with plasmid),positive control  group (transfected with pWPXLd),and experiment  group(transfected with pWPXLd-eGFP-of CD133);and the expression of eGFP gene in Hep-2 cells was observed under fluorescence microscope. Results  The specific 1 810 bp CD133 gene promoter  fragment was obtained successfully by PCR method. The eGFP  gene expression vector regulated by  human CD133 promoter was constructed successfully by PCR and enzyme digestion method.The  eGFP expressions in Hep-2 cells in  positive control group and experiment group were found,but weren’t in blank control group under fluorescence microscope. Conclusion The eGFP  gene expression vector regulated by  human CD133 gene promoter can regulate the expression of eGFP gene in laryngocarcinoma Hep-2 cells.

Key words: CD133 promoter, enhanced green fluorescent protein, tumor stem cell, Hep-2 cell line