Rotor-Gene Q MDx CE User Manual 02/2022
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6.
The Rel Min and Rel Max values are generated by calculating the standard deviation of the
quotient from the standard deviations of the GOI and Normalizer using the following formula:
2
2
cv
cv
cv
Norm
GOI
relconc
+
=
where:
meanvalue
stddev
X
s
cv
=
=
6.6.4
Delta delta C
T
relative quantitation
The delta delta CT method enables relative gene expression analysis. It is described by Livak and
Schmittgen (2001).*
This method does not require standard curves to be included in each run. Each sample is first
normalized for the amount of template added by comparison with the normalizing gene. These
normalized values are further normalized relative to a calibrator treatment. The calibrator could be,
for example, wild-type, untreated control, or time-zero samples.
It is essential that the amplification efficiencies of the gene of interest and the normalizing gene are
identical and that this is validated according to the guidelines of Livak and Schmittgen.
It is essential that the sample names are defined correctly in the
Edit Samples
window, with the
same samples labeled identically in each composite quantitation analysis.
1.
Analyze the data using “Quantitation”. It is not necessary to run a standard curve once
validation has been performed.
From the
Other
tab in the
Analysis
window, select
Delta Delta CT Relative Quantitation
. Select
New Analysis
.
* Livak, K.J. and Schmittgen, T.D. (2001) Analysis of relative gene expression data using real-time quantitative PCR and the
2^[–delta delta C(T)] method. Methods
25
, 402.