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. 2017 Nov 27;9(11):9618–9631. doi: 10.18632/oncotarget.22709

Figure 3. DNA methylation profile of the CpG island of TDRD9 promoter.

Figure 3

(A) Schematic diagram of the CpG island of TDRD9 promoter (-287 to +417 with respect to the transcription start site (TSS)). Amplified region for determination of DNA methylation (-234 to +109) is also shown. CpG pairs are represented by sticks. (B) Determination of DNA methylation by bisulfite conversion and sequencing of a region (-234 to +109 respect to the TSS) of the TDRD9 CpG island from different lung carcinoma cell lines. Open circles denote unmethylated CpGs, and filled circles represent methylated CpGs. Eight to fourteen independent clones of each sample were sequenced. (C) Relative mRNA expression levels of TDRD9 gene by RT-qPCR in TDRD9-negative lung carcinoma cell lines (H1264 and A549) after the treatment with 5-aza-2´-deoxycytidine for 72 hours. (D) Re-silencing of TDRD9 gene expression in the H1264 cell line following of 5-aza-2´-deoxycytidine removal. Cells were treated with 5-aza-2´-deoxycytidine for 72 hours and then washed and cultured in the absence of the drug. Cells were passed 1:3 dilution every three days, and samples were taken at the indicated passage (P) for RNA isolation. Data are the mean of at least n = 6 qPCR reactions from three independent experiments. Error bars represent ± SD values.