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《Chinese Journal of Clinical Medicine》 2006-02
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Construction and Identification of Recombinant Adenovirus Harboring the GIP-Human Insulin Gene

XU Meidong ZHANG Yiqun SHEN Kuntang,et al Department of General Surgery,Zhongshan Hospital,Fudan University,Shanghai 200032  
Objective:To construct a recombinant adenovirus harboring the human insulin gene regulated by the glucose-dependent insulinotropic polypeptide(GIP) promoter. Methods:GIP-hIns gene was cut from plasmid of pCDNA3-GIP-hIns and cloned into plasmid pCA13 - ACMV, which the plasmid pCA13 with the CMV promoter deletion and formed transfer plasmid of pCA13-GIP-hIns. The plasmid was co-transfected to 293 cells with plasmid pBHGE3 for package of recombinant adenovirus. The recombinant adenovirus vector was identified by PCR. The titer of the recombinant Ad was measured. A tumor-derived K-cell line STC-1 was transfected with Ad.GIP-hIns in vitro, RT-PCR was used to proved target gene expression in infected cells. Results: Positive recombinant adenovirus was obtained from 293 cells, and PCR test indicated that the recombinant Ad contained the GIP-hIns gene. The titer of purified Ad was 1×1011 pfu/ml. Meanwhile, mRNA product of hIns was detected in the infected cells by RT - PCR. Conclusion: A recombinant adenovirus harboring the GIP-hIns was successfully constructed, and it provides a good tool for further research on gene therapy Of diabetes.
【Fund】: 上海市科委基金资助课题(No:04DZ19505)
【CateGory Index】: Q78
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