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. Author manuscript; available in PMC: 2010 May 1.
Published in final edited form as: Biochim Biophys Acta. 2008 Nov 21;1787(5):290–295. doi: 10.1016/j.bbabio.2008.11.005

Fig. 2. Expression of Drosophila mtDNA helicase mutants analogous to human adPEO mutations in Schneider cells.

Fig. 2

Schneider cells with no plasmid (control) or carrying pMt/Hy (vector), pMt/WT/Hy (WT), pMt/W282L/Hy (W282L), pMt/R301Q/Hy (R301Q) or pMt/P302L/Hy (P302L) were cultured for 14 days in the presence or absence of 0.2 mM CuSO4. A, Protein extracts (20 μg) were fractionated by 9 % SDS-PAGE, transferred to nitrocellulose filters and probed with affinity-purified rabbit antiserum against d-mtDNA helicase or antiserum against d-ATPase β was indicated. B, Effect of overexpression of the deletion mutants on mtDNA copy number. Total mtDNA (10 μg) was extracted from Schneider cells (control) or Schneider cells carrying pMt/Hy (vector), pMt/WT/Hy WT ( ), pMt/W282L/Hy (W282L), pMt/R301Q/Hy (R301Q) or pMt/P302L/Hy (P302L) that were cultured for 14 days in the presence of 0.2 mM CuSO4. DNA was digested with XhoI, fractionated in a 0.7 % agarose/TBE gel, and then blotted to a nylon membrane. The membrane was hybridized with a radiolabeled probe for CytB, and then stripped and re-hybridized with radiolabeled probe for the histone gene cluster. The relative mtDNA copy number was quantitated as described under Methods.