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Chapter 3 Gene Expression and Other Quantitative Assays
Data Analysis
Data analysis varies depending on the product, assay, and instrument. Refer to the
appropriate instrument user guide for instructions on how to analyze your data.
General Process
The general process for analyzing the data from gene expression assays involves the
following procedures:
1. Viewing the amplification plots for the entire plate
2. Setting the baseline and threshold values
3. Using the methods detailed in this section to determine results
Resources for
Data Analysis
For more information about analyzing your data, see Livak and Schmittgen, 2001.
Also refer to the following documents:
• The appropriate instrument user guide
• User Bulletin #2: Relative Quantification of Gene Expression (PN 4303859)
• Data Analysis and Relative Quantification chapters in the TaqMan Cytokine
Gene Expression Plate I Protocol (PN 4306744). This protocol provides
examples using multiplex reactions.
• RQ Manager Software User Guide (PN 4339753)
• ABI PRISM® 7900HT Real Time PCR System User Guide (PN 4317596)
• SDS Enterprise Database Administration Guide (PN 4346513)
Note: Some documents are available through the Internet (see “How to Obtain
Support” on page xi).
Relative or Absolute Quantification
You can design your assay as relative or absolute quantification. How you calculate
the results of your Quantification Assays depends on your experimental design.
What Is Relative
Quantification?
Relative quantification describes the change in expression of the target gene in a test
sample relative to a calibrator sample. The calibrator sample can be an untreated
control or a sample at time zero in a time-course study (Livak and Schmittgen, 2001).
Relative quantification provides accurate comparison between the initial level of
template in each sample.
Example
You can compare the level of expression of a gene (Gene A) between a treated and
untreated sample (for example, a drug treatment) relative to the expression of a
second gene (Gene B). The resulting data is the expression of Gene A in treated and
untreated samples normalized to the expression of Gene B.
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DRAFT
November 13, 2003 12:42 pm, 3 QuantAssays.fm
Sequence Detection Systems Chemistry Guide