(1.Department of General Surgery, the Affiliated People′s Hospital of Jiangsu University, Zhenjiang Jiangsu 212002; 2.School of Medical Science and Laboratory Medicine, Jiangsu University, Zhenjiang Jiangsu 212013, China)
Abstract：Objective: To construct and identify recombinant adenovirus vector containing the human OX40-IgG1 fusion gene, then to infect the HL-7702 liver cells to filter out the best infective dose and detect expression of OX40Ig,which lay a foundation for the immune therapy inhibitory after organ transplantation.Methods: Specific OX40 and hIgG1 Fc primers were designed according to GenBank, and subcloned into the pAdTrack-CMV shuttle plasmid after PCR and identification. Retransformed into BJ5183 competent cells with pAdeasy-1 by calcium chloride method. The recombinant plasmid was detected by PCR and DNA sequence analysis and was transfected into 293A cells. The recombinant adenovirus infected HL-7702 liver cells and filtered out the best infective dose. OX40Ig gene expression was detected by indirect immunofluorescence assay. OX40Ig protein expression was detected by ELISA.Results:pTOX40Ig and pAdOX40Ig were constructed successfully according to restriction endonuclease analysis. 293A cells were transfected, and then the purified recombinant adenovirus infected HL-7702 liver cells.10 MOI was the best infective dose.OX40Ig effective expression was detected in HL-7702 liver cells. Conclusion: The recombinant adenovirus vector AdOX40Ig carrying the human OX40-IgG1 fusion gene was successfully constructed, and OX40Ig effective expression was found in HL-7702 liver cells in vitro.
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2013, Vol. 23 
