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. Author manuscript; available in PMC: 2009 May 1.
Published in final edited form as: Nat Struct Mol Biol. 2008 Oct 5;15(11):1169–1175. doi: 10.1038/nsmb.1499

Figure 2. Jhdm1b knockdown in primary MEFs inhibits cell proliferation and induces cellular senescence.

Figure 2

(a) Primary MEFs at passage 1 were transduced with lentiviral control (black bars), Jhdm1b (Jhdm1b KD, grey bars) or Ring1b (Ring1b KD, empty bars) shRNA and further selected by puromycin for 48 hours. mRNA levels were determined by quantitative real-time PCR (qRT-PCR). Samples were normalized against levels of Gapdh. The Jhdm1b or Ring1b expression level in empty vector control cells was arbitrarily set to 1. (b) Primary MEF cells transduced at passage 1 with lentiviral control (diamonds), Jhdm1b (Jhdm1b KD, squares), and Ring1b (Ring1b KD, triangles) shRNA and further selected by puromycin for 48 hours. 5×105 cells were plated and cell number was counted at different time points using a hemocytometer. (c) Primary MEFs were prepared as in (b), pulse labeled with BrdU and stained with PI and FITC conjugated anti-BrdU antibody. The percentage of BrdU positive cells was determined by flow cytometry for control (black bars), Jhdm1b KD (grey bars) and Ring1b KD (empty bars). (d) Cells prepared as in (b) were split at 1:5 for 5 passages and senescence associated β-galactosidase activity at pH 6.0 was determined by cellular staining. Data is presented for passage 1 (P1) and passage 5 (P5) cells. All error bars represent s.d. (n=3).