RG108 (N-Phthalyl-L-tryptophan) is an DNA methyltransferase inhibitor(IC50=115 nM).

DNA methyltransferase:115 nM

SM衍生的iPS细胞中,RG108能够明显降低DNA活性。在肿瘤细胞中,RG108抑制基因脱甲基和再活化,但是不影响着丝粒基因序列的甲基化。

In vitro methylation assay: The substrate DNA for the in vitro methylation assay is a 798 bp fragment (?423/+375 relative to the initiation codon) from the promoter region of the human p16Ink4a gene. The methylation reaction contains 350 to 400 ng substrate DNA and 4 units of M.SssI methylase (0.5 μM) in a final volume of 50 μL. Inhibitors are added to final concentrations of 10, 100, 200, and 500 μM, respectively. Reactions are done at 37 °C for 2 hours. After completion, the reaction is inactivated at 65 °C for 15 minutes and the DNA is purified using PCR Purification kit. Three hundred nanograms of purified DNA is digested for 3 hours at 60 °C with 30 units of BstUI and analyzed on 2% Tris-borate EDTA agarose gels.

For the determination of cellular growth and viability, cells are stained with trypan blue and counted using a standard counting grid.(Only for Reference)