Dual-Luciferase Reporter Assay for CPTH2 and siGCN5 Effects

The bim-luciferase (Luc) reporter, containing a 5.2 kb DNA fragment, was described previously.^{29 (link), 30 (link)} For the dual-luciferase reporter assays, cells were transfected with 1 μg of a luciferase reporter plasmid and 200 ng of the pRL–CMV Renilla luciferase reporter plasmid (Promega, Beijing, China). To test the effect of CPTH2 on the reporter, neurons were kept in conditioned media for 12 h after transfection and then transferred to serum-free media 25K containing CPTH2 (50 μM) for 24 h. To observe the effect of siGCN5s on the reporter, neurons were kept in conditioned media for 48 h after transfection. Firefly luciferase activity was normalized to Renilla luciferase activity as reported previously.^{37 (link)}