DNA Methylation Is Dispensable for Suppression of the Agouti viable yellow Controlling Element in Murine Embryonic Stem Cells
Figure 2
Analysis of IAP and agouti expression in Avy ES cells.
For qRT-PCR analyses, bar graphs represent mean+/− S.D. Results are representative of two independent experiments. A. qRT-PCR analysis of IAP expression. Expression levels are normalized to actin. RNA from Dnmt1c/c ES cells was used as a positive control for IAP expression. B. qRT-PCR analysis of IAP expression in cells either untreated or treated with the DNA demethylating agent 5-azacytidine (5-aza, 7.5 µM). Expression levels are normalized to actin and expressed as fold change relative to the signal observed in untreated cells. C. qRT-PCR analysis of agouti expression in the various organs indicated. Expression levels are normalized to GAPDH and measured relative to expression level in heart. D. qRT-PCR analysis of agouti expression in cells either untreated or treated with the DNA demethylating agent 5-aza. Expression levels are normalized to GAPDH and measured relative to untreated signals. E. qRT-PCR analysis of GFP expression in the various cell lines indicated. RNA from Oct4-GiP ES cells was used as a positive control for GFP expression. Expression levels are normalized to actin. F. Western blot data using an anti-GFP antibody on whole cell lysates from Avy/AGFP ES cells either untreated or treated with 5-aza or the histone demethylating agent Tricostatin A (TSA, 40 nM). Whole cell lysate from Oct4-GiP ES cells was used as a positive control. Western blot using an anti-tubulin antibody was used as a loading control.