Construction Of Recombinant P16 Adenovirus Vector And Its Suppressive Role In The Growth Of Human Heptocellular Carcinoma Cell Line

Objective To construct recombinant p16 adenovirus vector and explore the suppressive function of wild type p16 gene in the growth of human heptocellular carcinoma cell line.Methods Method to construct the recombinant p16 adenovirus vector, with the gateway cloning system, which was developed into a new technology in recent years, was applied. It is easy to transfer the DNA segment among vectors by the gateway-cloning system. The artificial synthesis sequence carrying sal I, Kozak consensus sequence for proper initiation of translation in mammalian cells, the three exons of the p16-INK4 suppressor gene from human and Ecol R, was transformed into competent cells. The sequencing of the artificial synthesis sequence was right. And pENTR1A-p16 vectors, the entry clones, were generated by a serial tests. Using gateway-cloning system, which the LR recombination reactions were finished, pAd/CMV/V5-DEST-p16 vectors, the expression clones, were created. The sequencing of the vectors was also right. The pAd/CMV/V5-DEST-p16 vectors digested by Pac I and purified were transfected into the 293A packaging cells by lipofection -mediated transfection. Western blot indicated that P16 protein was expressed in the infected SMMC7721. Growth of the tumor cells infected by Ad/p16 virus was inhibited.Results Recombinant adenovirus AdV/p16 could infect the hepatocellular carcinoma SMMC7721 cells and have the suppressive function in the growth of tumor cells.