The first commercially available software was the LightCycler® Relative Quantification Software (2001). It can be used to calculate and compare relative quantification results of triplicates of a target versus a calibrator gene. Target genes are corrected via a reference-gene expression and calculates on the basis of the median of the performed triplets. Real-time PCR efficiency correction is possible within the software and is calculated from the calibration curve slope, according to the established eq. 9, ranging from E = 1.0 (minimum value) to E = 2.0 (theoretical maximum and efficiency optimum). A given correction factor and a multiplication factor, which are provided in the product specific applications by Roche Molecular Biochemicals (LightCycler® Relative Quantification Software, 2001), have to be incorporated in eq. 6. Importantly, no statistical comparison of the results by a statistical test is possible.
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