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. 2011 Sep 28;6(9):e25634. doi: 10.1371/journal.pone.0025634

Figure 4. GSTP1 DNA methylation and protein expression in LNCaP cells after daily 5-aza-CdR treatment.

Figure 4

DNA and proteins were extracted from LNCaP cells treated with increasing doses of 5-aza-CdR (0.005–2.5 µM). Cells were treated daily and DNA and protein harvested after 6 days of treatment. (A) MSP was performed on bisulfite-modified DNA with primers targeting bisulfite-modified methylated GSTP1 promoter or unmethylated GSTP1 promoter. (B–C) the relative methylation status of the GSTP1 promoter following 5-aza-CdR treatment was further assessed by COBRA using two restriction enzymes, BstUI and HhaI. MDA-MB-231 breast cancer cells were used as a control for unmethylated GSTP1 promoter. (D) Immunoblot was performed to analyze GSTP1 protein expression. Detection of Hsp90 was used as a loading control.