关键词: 慢病毒载体, 癌症疫苗 , 粒细胞巨噬细胞集落刺激因子

Abstract: Objective: To construct a lentiviral vector encoding human granulocytemacrophage colony stimulating factor(hGM-CSF) genes and to transfect CT26 cells in order to establish the CT26 strains which has been continuously overexpressing the hGM-CSF gene. Methods: DNA fragments of human GM-CSF were amplified by PCR and were inserted into the vector plasmid L166. The three plasmids expressing lentivirus were packaged into virus packaging cell line 293T using the CaCl2 method. After 72 hours of transfection, the virusproducing cell supernatant was produced. CT26 cells were transfected by lentivirus and the titration was determined. Results： The sequence of the PCR product in the recombinant plasmid was produced. The three plasmids were effectively transferred into 293T. When the CT26 cells were infected as target cells by the lentivirus supernatant, the titration of lentivirus was 4.69×105IU/ml. hGM-CSF was determined in the supernatant of CT26 cells transfected by lentivirus, which was kept for more than two months. The transfected CT26 cells were screened by puromycin at a terminal concentration of 15μg/ml. Conclusion： The lentivirus encoding hGMCSF genes were successfully constructed.

Key words: Granulocytemacrophage colonystimulating factor, Lentivirus vector, Cancer vaccines