Bio-Rad Firmware & Software Updates - Manual (Page 101)

Gene Expression Analysis
90
• Relative expression level of one target sequence in one sample compared to the
same target under different sample treatments. For example, the relative amount of
one gene relative to itself under different temporal, geographical, or developmental
conditions
Plate Setup for Gene Expression Analysis
To perform gene expression analysis, the contents of the wells must include the following:
• Two or more targets. The two targets that represent different amplified genes or
sequences in your samples
• One or more reference targets. At least one target must be a reference target for
normalized expression. Assign all reference targets in the Experiment Settings window
(page 48) to analyze the data in Normalized Expression mode (C
q
). Runs that do not
contain a reference must be analyzed using Relative Expression mode (C
q
)
• Common samples. Your reactions must include common samples (minimum of two
required) to view your data plotted in the Gene Expression tab. These samples represent
different treatments or conditions for each of your target sequences. Assign a control
sample (optional) in the Experiment Settings window (page 48)
The requirements for Gene Expression setup in the Plate Editor depend on whether reaction
contents are singleplex PCR with one fluorophore in the reactions, or multiplex PCR with
more than one fluorophore in the reactions.
Figure 64 shows an example of the minimum contents of the wells for a singleplex gene
expression run.
Figure 64. Example of well contents in a singleplex gene expression run.
Figure 65 shows an example of the minimum contents of the wells for a multiplex gene
expression run.
Figure 65. Example of well contents in a multiplex gene expression run.
Guided Plate Setup
If the plate setup of a data file does not contain the required information for analysis and the
Gene Expression tab is selected, the space normally occupied by the bar chart will contain
instructions for entering this information. For normalized gene expression, the following steps
must be completed:
1. Define Target and Sample names using
• Plate Setup. This will open the Plate Editor window
• Replace Plate File. Select a plate layout file that was previously created