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. 2010 Sep 13;39(1):104–118. doi: 10.1093/nar/gkq774

Figure 1.

Figure 1.

Generation of cell lines stably overexpressing DNA methyltransferases (DNMTs). (A) Schematic structure of DNMT isoforms and expression vector constructs. The conserved PWWP and PHD-like domains, DNMT catalytic motifs (I, IV, VI, IX and X), and alternative splicing sites are indicated. The coding regions of DNMT 3B4 and DNMT 3B5 in red represent frame shift mutations generated by alternative splicing. The expression vectors encode N-terminal myc-tagged DNMT isoforms. (B) mRNA expression of exogenous DNMTs. mRNA expression of each DNMT isoform was assessed by real time RT-PCR with primers designed for the IRES region located on a bicistronic transcript and normalized to the expression of GAPDH. (C) Protein expression levels of exogenous DNMT isoforms by western blotting using a myc antibody. The expression level of DNMTs shown in the right panel was lower than those in the left panel, such that a 10-fold higher amount of protein was loaded in the left panel, as shown by the GAPDH control. Note the 10-fold higher amount of DNMT 3A1 in the left panel was compared with 3A1 (1/10) in the right panel.