| Objectiver: To investigate the possibility of gene immunization with macaca Rhesus chorionic-gonadotropin hormone (rmCG) β subunit encoding gene in inducing specific anti-tumor immunity, and to seek for a new pathway of immunity biotherapy of tumor.Methods: The synthesized rmCG β encoding gene , amplified by PCR and modified by restriction endonucleases digestion , was subcloned into an eukaryotic expression vector pcDNA3.1(+) and a recombinant eukaryotic expression vector pcDNA3.1(+)-Sig-Pr~+-IgGFc-GPI to construct pcDNA3.1(+)-rmCG β and pcDNA3.1(+)-rmCG β -Pr~+-IgGFc-GPI plasmid. The plasmids were identified by PCR and restriction endonucleases digestion as well as DNA sequencing. Protective assay was performed by subcutaneous inoculation of Renca cells into the immunized mice. The weight of the tumor was evaluated after challenge. We constructed the eukaryotic expression vector pcDNA3.1(+)-Sig-Pr~+-IgGFc-GPI,pcDNA3.1(+)-rmCG β -Pr~+-IgGFc-GPI and transfected pcDNA3.1(+)-rmCG β -Pr~+-IgGFc-GPI into the B16 cells by means of liposome-mediated transfection method, and then the resultant positive clone was multiplied in culture. The transfection was confirmed to be successful by RT-PCR. BALB/c mice were immunized i. m. with 20μg, 40μg, 100μg DNA of pcDNA3.1(+)-rmCGβ plasmid, 40μg DNA of pcDNA3.1(+)-rmCGβ -Pr~+-IgGFc-GPI plasmid and mock DNA , respectively. BALB/c mice were divided into rmCG and control groups, which were injected through the tail vein, respectively with the pcDNA3.1 (+) -rmCG-Pr~+-IgGFC-GPI transfected and pcDNA3.1 (+) transfected B16 cells, and examined for the number of metastatic tumor focus in lung after two weeks. |
