Mitochondrial DNA quantification.

Total DNA was extracted from H9c2 cells after 48 h of transfection with miRIDIAN microRNA Mimics, rno-miR-140-5p mimic (Dharmacon, No. C-320700-00), or Mimic Negative (Scramble) Control (Dharmacon, No. CN-001000-01) with DNeasy Blood & Tissue Kit (Qiagen, No. 69504) following manufacturer's protocol. Mitochondrial DNA (mtDNA) copy number relative to nuclear DNA (nucDNA) copy number was quantified by qPCR using the equation: relative mtDNA content = 2 × 2^ΔCT, where CT = (nucDNA CT − mtDNA CT) as described previously (53). To quantify the amount of mtDNA, the primer set located within the mitochondrial 16s rRNA region was used, which are 5′-TGTGGAATTAGTTGTGTGTGTAAG-3′ Tm=59 and 5′-AGGTGAAAAGCCTATCGACCTTG-3′ Tm = 62. To quantify the amount of nuclear DNA, the primer set located within the actin region was used, which are 5′-GCGGTGACCATAGCCCTCTTT-3′ Tm=61 and 5′-TGCCACTCCCAAAGTAAAGGGTCA-3′ Tm=62.