Comparison of expression cassettes with modification or deletion of WPRE. A. Schematic drawings of expression cassettes tested. Each EGFP expression cassette was transduced into hippocampal neuron cultures together with tdTomato expression cassette, which contains complete WPRE and bGHpA. (ITR, Inverted Terminal Repeat; CaMKIIp, CaMKIIα promoter; WPRE, Woodchuck hepatitis virus posttranscriptional regulatory element, bGHpA: bovine growth hormone polyadenylation signal, INT: chimeric intron) B-C. Representative images of cell fluorescence (B), and western blot analysis of EGFP and tdTomato expression (C). Scale bar, 200 μm. D. Summary graph of western blot analysis shows that the expression level of EGFP driven by CW3B having WPRE3 (247 bp) is comparable (83.4%) to CWB having WPRE (600 bp). In contrast, EGFP expression level was significantly reduced in CIB (16.6%, P < 0.001) and CB (28.6%, P < 0.001) compared to CWB. (***P < 0.001, ANOVA Bonferroni’s post-hoc comparison).