The Structure And Expression Of Adenoviruses Carrying11R-P53and MGM-CSF And It Inbitionaleffect Towards Tumor Tissue

Immunotherapy and genetherapy of the hepatocellular carcinoma has currently shown potential effect and good future. Antioncogene P53and gene GM-CSF was recombined into the same adenovirus vector in this text. Before large-scale preparation of plasmid DNA, recombined adenovirus vector was transfect into packaging cell. The recombinant gene was renamed as SG655-mGMP. The potential anti-tumor effect was identified by testing the expression of desired gene in vitro and in vivo. SG655-mGMP was expect to show potent effect towards tumor cell which can kill or stop the growth of the tumor cell through gene level and cell immunology level.Aim:To construct an adenoviral vector carrying11R-P53and mGM-CSF gene and also detect its expression in tumor cell. Method. Recombination these genes to the area of E1B-55KD by combined bisulfite restr iction assay. Adenovirus packaging and prepare plasmid in quantity after DNA sequencing, and named as SG655-Mgmp. Virus titer was tested by TCID50, and transfection in vitro was conducted to293cells, Huh7cells and Hepal-6cells using SG655-mGMP(MOI=5) and then teste the expression of the desired gene by western blot and ELESA in vivo. Forty C57BL/6mouse was injected with Hepal-6during logarithmic phase,5x106cells per mouse. The mouse were distributed into four groups named A,B,C and D, and was injected with SG655-mGMP, SG7605-11R-P53, SG7605-mGM-CSF and normal saline (NS). The expression of the desired gene was testing by Immunohistochemistry. Result. The SG655-mGMP was structured successfully and the titer is7.3×109PFU/ml. And the desired gene can express in vivo and in vitro.