Depletion of murine Dnmt proteins after 5-Aza-CdR treatment.

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Depletion of murine Dnmt proteins after 5-Aza-CdR treatment. Depletion of murine Dnmt proteins after 5-Aza-CdR treatment. A. Dnmt protein levels in wild-type (M1/3A/3B; WT) ES cells and Dnmt1−/− (3A/3B), 3b−/− (M1/3A), 3a−/− (M1/3B), and 3a/3b−/− (M1) knockout ES cells. Equal amounts (30 μg) of protein extracts were analyzed by SDS-PAGE, and Dnmt proteins were identified using antibodies specific for each Dnmt protein. The two bands identified by the Dnmt3b antibody represent the alternatively spliced variants Dnmt3b1 and 3b6. B. Western blot of Dnmt protein levels in mouse ES cells under untreated conditions or after treatment with 5-Aza-CdR. Protein cell lysates were collected from untreated and 5-Aza-CdR-treated cells 24 h after the initial drug treatment. Expression levels of β-actin were determined to confirm accurate protein loading. C. Semiquantitative RT-PCR of Dnmt mRNA levels in ES cells under untreated or treated conditions. H4F2 was used as a marker for cell proliferation. Daniel J. Weisenberger et al. Mol Cancer Res 2004;2:62-72 ©2004 by American Association for Cancer Research