Constructing a lentiviral vector overexpressing indoleamine 2,3-dioxygenase / 中国组织工程研究

ABSTRACT

BACKGROUND:Immune rejections after organ transplantation or serious adverse reactions due to immunosuppressive drugs show a lack of effective treatments and poor therapeutic outcomes. Therefore, we try to find an effective immune suprresion method in combination of the latest immunomodulatory achievements. OBJECTIVE:To construct a lentiviral vector overexpressing indoleamine 2,3-dioxygenase (IDO). METHODS:(1) The IDO gene that was successful y contructed was inserted into lentiviral packaging plasmids GV308 to construct GV308-IDO lentivirus packaging plasmids. (2) The 293T cel s with 80%confluence were co-cultured with 5'LTR and 3'LTR, basic elements of lentiviral packaging auxiliary components, including Psi, cPPT, 3FLAG, TetR, IRES, WRPE, TetIIP, Ubiquitin Promoter, SV40 origin and HIV. RESULTS AND CONCLUSION:Western blot assay showed that in 10 g/L agarose gel electrophoresis, there was a target fragment at Mr 48 000. This value was consistent with the size of IDO protein. RT-PCR results showed visible IDO expression in 293T cel s. These findings suggest that IDO fusion gene has been successful y reorganized in the lentiviral packaging plasmids.