Levels of plasma circulating cell free nuclear and mitochondrial DNA as potential biomarkers for breast tumors

Table 1 Quantitative PCR (qPCR) for GAPDH and MTATP8

Gene	Gene ID	Sequences of primers and probes (5' → 3')		Length of primer/probe	Amplicon lengths (bp)
GAPDH	2597	Forward	CCC CAC ACA CAT GCA CTT ACC	21	97
		Reverse	CCT AGT CCC AGG GCT TTG ATT	21
		Probe	(MGB) TAG GAA GGA CAG GCA AC (VIC)	17
MTATP8	4509	Forward	AAT ATT AAA CAC AAA CTA CCA CCT ACC	27	78
		Reverse	TGG TTC TCA GGG TTT GTT ATA	21
		Probe	(MGB) CCT CAC CAA AGC CCA TA (FAM)	17

Quantitative PCR (qPCR) for GAPDH and MTATP8 was carried out in a total reaction volume of 25 μl containing 7 μl H₂O, 12.5 μl TaqMan^® Universal PCR Master Mix, 0.75 μl of each of the shown 10 μM primers, 1 μl of a 5 μM FAM-labeled MTATP 8-probe and 0.5 μl of a 5 μM VIC-labeled GAPDH-probe and 1 μL of template. The reaction was performed at the following conditions: initiation for 2 minutes at 50°C, followed by a first denaturation for 10 minutes at 95°C and a further step consisting of 40 cycles of 15 seconds at 95°C and 1 minute at 60°C.

ISSN: 1476-4598