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. 2017 Jun 6;45(14):8524–8540. doi: 10.1093/nar/gkx508

Figure 1.

Figure 1.

RBM10 overexpression (OE) downregulates endogenous RBM10 RNA and protein expression. (A) Relative levels of 5’-untranslated region (5’UTR) and 3’UTR derived from endogenous RBM10 mRNA in the absence (Ctrl) and presence of overexpressed RBM10 (RBM10 OE) in HEK293 cells. Expression levels were estimated using RNA-Seq data and normalized with respect to GAPDH expression (n = 2; error bar: range). (B) RT-PCR analysis of total and endogenous RBM10 mRNA expression in tet-on HEK293 cells inducibly expressing RBM10-EGFP in response to doxycycline (Dox), using primers designed to amplify RBM10 coding region (CDS), 5’UTR and 3’UTR sequences, respectively. Left panel: representative agarose gel images of RT-PCR products. Right panel: qPCR results. Data are presented as mean ± SEM for n = 4 biological replicates. **P < 0.01, ***P < 0.001 (one-way ANOVA followed by Dunnett's tests). (C) Western blot analysis of RBM10 protein expression under conditions in (B), using an antibody that recognizes both endogenous and overexpressed RBM10-EGFP. α-Tubulin served as a loading control. A representative result from three independent experiments is shown. The faint band above the endogenous RBM10 band in RBM10-EGFP overexpressed samples, indicated by the open arrowhead, is likely an artifact resulting from RBM10-EGFP overexpression (see also Supplementary Figure S1D).