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Analysis of promoter methylation, mRNA and protein expression of claudin-11 in gastric cell lines.

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posted on 21.02.2013, 03:44 by Rachana Agarwal, Yuriko Mori, Yulan Cheng, Zhe Jin, Alexandru V. Olaru, James P. Hamilton, Stefan David, Florin M. Selaru, Jian Yang, John M. Abraham, Elizabeth Montgomery, Patrice J. Morin, Stephen J. Meltzer

This figure illustrates the claudin-11 promoter methylation, mRNA expression levels and protein expression in gastric cells lines. A) Quantitative methylation-specific PCR (qMSP) for CLDN11. Genomic DNAs isolated from immortalized human normal gastric epithelial cells (HFE145) and GC cell lines AGS, SIIA, MKN28, KATOIII, and SNU-1 obtained from ATCC were analyzed by qMSP. This Figure illustrates that the promoter region of CLDN11 gene is hypermethylated in all GC cell lines relative to HFE145 cells. B) CLDN11 mRNA expression in gastric cell lines. Total RNAs from different gastric cell lines were subjected to quantitative real-time RT-PCR analysis. As can be seen in this figure, HFE145 cells expressed very high levels of CLDN11 mRNA, while all five cancer cell lines tested had very low or undetectable CLDN11 mRNA expression. C) Western blot analysis of claudin-11 expression in gastric cell lines. Total cell lysates obtained from various gastric cell lines were probed with the anti-claudin-11 antibody. This figure illustrates that while the immortalized normal gastric epithelial cell line, HFE145, expressed abundant claudin-11 protein, it could not be detected in various GC cell lines. Anti-β-actin antibody was used as a loading control.

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