Figure 4. Effect of targeting siRNAs suppressed TβRII protein and mRNA expression by western blot and RT–PCR. A: siRNAs suppress TβRII protein expression in SRA01/04 cells. Total cell lysates from SRA01/04 cells treated with hT1, hT2, and hT3 siRNA or control, scrambled siRNA for 48 h were separated on 12% SDS polyacrylamide gels and immunoblotted with a TβRII specific antibody. Lane 5 contains lysate from cells incubated with Lipofectamine™ 2000 reagent alone (no siRNA). Lane 4 contains lysate from cells treated with 80 nM scrambled siRNA. Lanes 1, 2, and 3 contain lysates of cells treated with hT1,hT2, and hT3 siRNA at a final concentration of 80 nM. The TβRII band is indicated. B: siRNA reduces levels of TβRII transcript. RNA was prepared from cells treated with scrambled siRNA or hT 3 siRNA for 48 h at 80 nM/ concentration. Following conversion to cDNA, the samples were analyzed by real time PCR using primers to TβRII. The bar graphs show the mean expression levels of TβRII transcript normalized to ACTB relative to that of scrambled siRNA treated controls. Asterisks indicate that the data are significantly different from the scrambled controls (p<0.0001, n=3, one-way ANOVA, Dunnett-test).