Effects of SOX2 overexpression on CD133

<p>⁻<b>DAOY cells.</b> i-SOX2-DAOY cells were FACS sorted into CD133⁻ and CD133⁺ populations. SSEA1⁺ cells were excluded from both populations. (A) CD133⁻ cells were cultured in medium without or with Dox (0.5 µg/mL) for 24 hours. Cells were then collected for FACS analysis of surface expression of CD133 and SSEA1. (B) RNA was isolated from CD133⁻ cells cultured without and with Dox (0.5 µg/mL) for 24 hours. cDNA was prepared and RT-qPCR was conducted. Values represented are differences in Ct value (normalized as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0044087#pone-0044087-g007" target="_blank">Figure 7</a>), where a positive value indicates increased expression of a given transcript within the Dox supplemented population. (C) A scatter plot of transcript expression, as determined by Affymetrix microarray analysis, comparing i-SOX2-DAOY cells cultured in the absence or presence of Dox (0.5 µg/mL) for 24 hours. Transcript expression from CD133⁻ DAOY cells cultured without Dox is represented on the y-axis; whereas, transcript expression from cells cultured with Dox is represented on the x-axis. A single replicate of each condition was analyzed. RT-qPCR analysis (subfigure B) was used to validate the array data. (D) Gene ontology analysis comparing genes that were upregulated or genes that were downregulated 2-fold, in CD133⁻ DAOY cells cultured in the absence or presence of Dox. The y-axis represents the percentage of up- or downregulated transcripts that reside within various ontology categories.