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. 2013 Jan 25;41(5):3144–3161. doi: 10.1093/nar/gkt029

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© The Author(s) 2013. Published by Oxford University Press.

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 8. — Overexpression of Ddk1 decreases 7S DNA levels and has no substantial effect on mtDNA levels. (A) Subcellular localization of Ddk1-FLAG in a stable inducible 293 cell line. Exogenous gene expression was induced for 24 h, and cells were subjected to immunoﬂuorescence staining and confocal microscopy. The same microscope settings were used for uninduced and induced cells. Mitochondria were labelled with MitoTracker, and Ddk1-FLAG with primary mouse anti-FLAG antibodies followed by detection with secondary AlexaFluor 488 conjugated antibodies. Nuclei were stained with Hoechst dye. The bar represents 10 μm. (B) Western blot analysis of Ddk1-FLAG level in parental 293 or transgenic cell lines. Expression of the Ddk1-FLAG was induced for the indicated times. Samples from two independent induced cultures were analysed. Ponceau S staining of the membrane was performed for standardization. (C and D) Analysis of mtDNA (C) and 7S DNA (D) levels in parental 293 cells and their derivative that overexpresses (or not) Ddk1 for the indicated time. Levels were examined using Southern blots. Graphs present the mean values obtained in two independent experiments. Error bars represent standard deviation. The signal arising from hybridization to the nuclear 28rDNA gene (nDNA) was used as a loading control. (C) Differences are not statistically significant (ANOVA test). (D) The P-values obtained in an ANOVA test are indicated. ns, statistically insignificant; P > 0.05.