Construction of TK/RFP Fusion Gene Expression Vector and Its Expression in HepG2 Cells

Objective: To construct gene transfer system for red fluorescent protein(RFP)marked herpes simplex virus thymidine kinase (HSV-TK) gene into eukaryotic and to investigate its expression in HepG2 cells. Methods: HSV-TK gene was amplified from pBLuescript-TK by PCR and inserted into plasmid pDsRed2-N1. Using lipofectin method, the recombinant expression plasmid pDsRed2-N1-TK was transfected into HepG2 cells. The expressed mRNA of TK gene was detect by RT-PCR, and the RFP expression was observed under fluorescence microscope. Results: When detected by fluorescence microscope, the red fluorescence was localized in the nucleus of HepG2 cells of test group, and in the cytoplasm of control group; TK gene mRNA was detected in the transfected cells. The expression of recombined plasmid pDsRed2-N1-TK in HepG2 cells was proved by RT-PCR. Conclusions: The HSV-TK and RFP fusion gene can express in HepG2 cells. The expressed fusion protein show double activity of HSV-TK and RFP; RFP can be used as a reporting gene for tracing the expression of HSV-TK gene, which provide practical and feasible tools for the study of gene therapy to cancer using herpes simplex type thymidine kinase gene transfer.