Protocol detail

Mitochondrial DNA Copy Number Quantification

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mtDNA copy number was determined as described, by calculating the ratio of mtDNA to nuclear DNA (ACTB) per cell in relation to standard curves of known concentrations, ranging from 10⁻¹ ng/μL to 10⁻⁸ ng/μL [42 (link)]. Quantitative PCR was performed on a Rotor Gene RG-3000 (Corbett Research, Mortlake, Australia), with RotorGene software (v6.0) used for data acquisition and analysis. Standard curves were used with a correlation coefficient of R² > 0.9 and an efficiency coefficient of R > 0.95. Mitochondrial DNA copy number per cell was calculated as follows:

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Burgin H.J., Crameri J.J., Stojanovski D., Sanchez M.I., Ziemann M, & McKenzie M. (2022). Stimulating Mitochondrial Biogenesis with Deoxyribonucleosides Increases Functional Capacity in ECHS1-Deficient Cells. International Journal of Molecular Sciences, 23(20), 12610.

Publication 2022

Rotor Gene RG-3000 RotorGene software

Cell Gene Mitochondrial dna

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Variable analysis

independent variables

Concentrations of mtDNA and nuclear DNA (ACTB) standards, ranging from 10^-1 ng/μL to 10^-8 ng/μL

dependent variables

Ratio of mtDNA to nuclear DNA (ACTB) per cell

control variables

Quantitative PCR performed on a Rotor Gene RG-3000 (Corbett Research, Mortlake, Australia)
RotorGene software (v6.0) used for data acquisition and analysis
Standard curves with a correlation coefficient of R^2 > 0.9 and an efficiency coefficient of R > 0.95

controls

Positive control: Standard curves of known concentrations of mtDNA and nuclear DNA (ACTB), ranging from 10^-1 ng/μL to 10^-8 ng/μL
Negative control: Not explicitly mentioned

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