Construction of sense and antisense human heparanase and green fluorescent protein eukaryotic coª²expressing vectors and their expression in lung cancer cell lines

    NIU Huiª²Jun, YANG Shiª²Ming, FAN Shiª²Zhi, CAI Yongª²Guo, FANG Dianª²Chun, LUO Yuanª²Hui, WANG Dongª²Xu

    1Department of Gastroenterology, Southwest Hospital, Third Military Medical University, Chongqing 400038, China, 2Department of Cardiothoracic Surgery, Daping Hospital, Third Military Medical University, Chongqing 400042, China

    ¡¾Abstract¡¿ AIM: To construct sense and antisense human heparanase and enhanced green fluorescent protein eukaryotic expressing vectors and to transfect these vectors into lung cancer cell lines. METHODS: The human Heparanase cDNA fragment contained in the pcDNA3ª²Hpa vector was cloned into the enhanced green fluorescent protein eukaryotic expressing vector pIRES2ª²EGFP in cisª²direction or transª²direction. The recombinant vectors were further identified by digestion of BamH I and transfected to lung cancer cells by liposome method. After selection with G418, the transfected efficiency was observed under fluorescent inverted microscope. RESULTS: After digested by BamH I, two fragments with the length of 5.3 and 1.7 kb, respectively, formed in sense fluorescent eukaryotic expressing vector (pIRES2ª²EGFPª²sHpa), while two other fragments with the length of 6.5 kb and 0.5 kb, respectively, formed in antisense fluorescent vector (pIRES2ª²EGFPª²aHpa). Electrophoretic results were completely identical with the theoretical calculation. Green fluorescence of the transfected cells was observed under fluorescent inverted microscope. CONCLUSION: Human heparanase sense and antisense fluorescent eukaryotic expressing vectors are successfully constructed and transfected to lung cancer cells. ......