Figure 7

Quantitative real-time RT-PCR to validate the results of microarray analysis. Results of quantitative real-time RT-PCR for 3 (CXXC5, RAB20, LHX8) out of 7 seven genes studied are shown here. These genes were found to be significantly different (P > 0.001) by microarray analysis in Lenti 1+3 experimental group, in which both ALDH1A1 and ALDH3A1 were knocked down by siRNA expression, in comparison to WT control group. The real-time RT-PCR was performed on RNA obtained from 5 cell lines including cells that express siRNA against ALDH1A1 only (Lenti 1), against ALDH3A1 only (Lenti 3), or against green fluorescent protein (GFP) as another control, in addition to the Lenti 1+3 and WT groups. The results are expressed as the log 10 of relative change in each gene expression with the denominator being the 18S as a reference control and using the fold increase (+) or decrease (-) compared to the WT/18S ratio as the baseline expression value. The direction (up- or down-regulation) for all 7 genes confirmed the results of microarray analysis even in the Lenti 1 and Lenti 3 cells. LHX8 was the only gene out of the 7 tested that was non-specifically down-regulated since it showed similar changes in the GFP control group as well.