Transfection and Luciferase Assay Protocol

Cotransfections of siRNA and CXCL10-Luc promoter constructs were done in 24 wells using 7.5 pmol siRNA, 62.5 ng luciferase reporter plasmid and Lipofectamine 2000 (Invitrogen) according to the manufacturer's instruction. 48 h later the cells were harvested in 100 mM KPO₄, pH 7.8, 0.1% NP40, and 1 mM DTT and the luciferase activity was determined with Promega's GloMax® Multi Reader as described previously [12 (link)]. RTS3b cells were transfected with the appropriate expression vectors using Fugene HD (Roche Diagnostics). The preparation of cell extracts, Western Blots (WB), and coimmunoprecipitations were performed as described previously [12 (link)].