Abstract: Objective To construct the adenoviral overexpressed vector of Krüppel-like factor 9(KLF9) and to explore the role of KLF9 in the regulation of anti-ROS genes expression. Methods The KLF9 gene is first cloned into the shuttle vector pAdTrack-CMV. The resultant plasmid is linearized by digesting with restriction endonuclease Pme I, and subsequently cotransformed into E. coli. BJ5183 competent cells with pAdEasy-1.Recombinants are selected for kanamycin resistance and recombination confirmed by restriction endonuclease analyses. Finally, the linearized recombinant plasmid is transfected into 293A cells. Recombinant adenoviruses are typically generated within 7 to 12 days. The mouse primary hepatocytes are isolated from the C57BL/6J mouse and subsequently H2O2 treatment is conducted. We aslo performed KLF9 overexpression by using Ad-virus system (Ad-KLF9) in mouse primary hepatocytes isolated from.Quantitative real-time PCR (qRT-PCR) analysis was further performed to determine the expression of anti-ROS genes including catalase(CAT), manganese superoxide dismutase(SOD2) and genes were upregualted in C57BL/6J mouse primary hepatocytes following H2O2 treatment (P<0.05).The results of qRT-PCR and western blot replied that adenoviral vector of KLF9 was successfully produced with 90% infection efficiency. The expression of anti-ROS genes were also upregualted in C57BL/6J mouse primary hepatocytes after infecting with Ad-KLF9(P<0.05).Conclusion KLF9 represents its impact on the expression of anti-ROS genes.

Key words: Key words: KLF9, Overexpression, Adenovirus, ROS