Chapter 6 Design the Comparative C
T
Experiment
Define the Methods and Materials
Applied Biosystems 7500/7500 Fast Real-Time PCR System Getting Started Guide for Relative Standard Curve
and Comparative C
T
Experiments
118
Notes
–Select RNA if you are performing 1-step RT-PCR. You are adding total RNA or
mRNA to the PCR reactions.
Note: To use the Fast ramp speed with RNA templates, you must design your
experiment using Advanced Setup instead of the Design Wizard.
–Select gDNA (genomic DNA) if you have already extracted the gDNA from
tissue or sample. You are adding purified genomic DNA to the PCR reactions.
For More
Information
For more information on:
• Completing the Methods and Materials screen – Open the 7500 Software Help by
clicking or pressing F1.
• Determining PCR efficiencies – Open the 7500 Software Help by clicking or
pressing F1. Within the Help, search as follows:
a. Select the Search tab.
b. Enter PCR efficiency.
c. Click List Topics.
d. Double-click Determine Amplification Efficiency.
• Using Advanced Setup – See “Advanced Setup Workflow” on page 204.
• Using the relative standard curve quantitation method – See Chapters 2 through 5 of
this guide.
• Using the standard curve quantitation method – Refer to the Applied Biosystems
7500/7500 Fast Real-Time PCR System Getting Started Guide for Standard Curve
Experiments.
• TaqMan and SYBR Green reagents – Refer to the Real Time PCR System Reagent
Guide.
• PCR, including singleplex vs. multiplex PCR and 1-step vs. 2-step RT PCR – Refer
to the Real Time PCR System Reagent Guide.
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