| Popis: |
Additional file 5: Figure S4 (part 2). Related to Figs. 3–7. CRISPRa is highly specific in hit-and-run and lentiviral approaches in HCC cell lines. A-K The potential off-target genes (MMP11, MOCOS, PIR, SEMA5A, TMEM14C, HRH1, NACC2, NONO, PPA1, RBM39, NKAIN3, and ABCC3) found in proximity to genomic regulatory regions for the most potent gRNAs utilized were assessed by qRT-PCR 48 h post-transfection, employing SpdCas9-VPR and MS2-MCP-p65-HSF1 system in HuH-7 (A), (B), (C), (E) and Hep3B cells (F), (H), (J); and in stable Hep3B cell lines expressing SpdCas9-VPR (D), (G), (I), (K). Relative gene expression, expressed as fold change, was normalized to cells transfected with empty vector control (EV) or to cells transduced with SpdCas9-VPR NO G, and compared to either EV or NO G conditions for statistical analysis. Data presented as means ± SEM (n = 3), and P-values were determined by two-way ANOVA with Dunnett's multiple comparisons test for panels (A), (B), (C), (E), (F), (H), and (J) (A: ***P = 0.0001; B, C, E, F, and H: ****P < 0.0001; J: ***P = 0.0003, ****P < 0.0001); or by two-way ANOVA with Šídák's multiple comparisons test for panels D, G, I, and K (****P < 0.0001). |
