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Analyzing real-time PCR data by the comparative CT method


Two different methods of presenting quantitative gene expression exist: absolute and relative quantification. Absolute quantification calculates the copy number of the gene usually by relating the PCR signal to a standard curve. Relative gene expression presents the data of the gene of interest relative to some calibrator or internal control gene. A widely used method to present relative gene expression is the comparative CT method also referred to as the 2 - ΔΔ C T method. This protocol provides an overview of the comparative CT method for quantitative gene expression studies. Also presented here are various examples to present quantitative gene expression data using this method.

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Figure 1: Real-time PCR output, calculation of CT.
Figure 2: Real-time PCR plots from gene expression profiling.
Figure 3: Real-time PCR expression profiling of microRNA in pancreas tissue.


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Correspondence to Thomas D Schmittgen.

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Schmittgen, T., Livak, K. Analyzing real-time PCR data by the comparative CT method. Nat Protoc 3, 1101–1108 (2008).

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