Fig. 5.

Resolution of the assay. a Amplification curves obtained with control primer pair 1 (dashed lines) or control primer pair 2 (solid lines) and different copy numbers of both templates present in equimolar ratio shown as means of triplicates performed in independent experiments with similar results (n = 5). The corresponding standard curves are shown on the right. Correlation coefficient (R ²), slope (M), y-intercept (B) and amplification efficiencies (E) calculated with the equation E = 10^−1/M are indicated. b Copies obtained in different reactions (x) with control primer pair 1 or control primer pair 2 at different template concentrations were calculated using the equation: $\text{copies}=\left[{\text{N}}_0\left(\text{X}\right)/{\text{N}}_0\left(\text{con2}\right)\right]·{10}^8=2^{\text{Cqz}·log\left(\text{Ez}\right)/log\left(2\right)\text{-}\text{Cqx}·log\left(\text{Ex}\right)/log\left(2\right)}·{10}^8$ with Cqz and Ez = Cq and E values obtained with control primer pair 2 and 10⁸ introduced copies. The efficiencies introduced into the equation were obtained either from the comparative quantitation tool of the Rotorgene software (red columns) or the standard curves shown in A (blue columns). Dark columns represent reactions using control primer pair 2, light columns control primer 1. Mean values ± SE of five independent experiments each performed in triplicate are presented