Chapter 6 Design the Comparative CT ExperimentDefine the Methods and MaterialsApplied Biosystems 7500/7500 Fast Real-Time PCR System Getting Started Guide for Relative Standard Curveand Comparative CT Experiments116NotesDefine the Methods and MaterialsIn the Methods and Materials screen, select the quantitation method, reagents, rampspeed, and PCR template to use for the experiment.About theExampleExperimentIn the comparative CT example experiment:• The comparative CT quantitation method is used.• TaqMan ® reagents are used.• The standard ramp speed is used in the instrument run.• cDNA (prepared from total RNA isolated from liver, kidney, brain, and universaltissues) is the template type. You must first perform reverse transcription to convertthe RNA to cDNA (see “Prepare the Template” on page 139).Complete theMethods andMaterials Screen1. Select Comparative CT (∆∆CT) as the quantitation method.2. Select TaqMan ® Reagents for the reagents. (default)IMPORTANT! Fast runs cannot be performed on a 7500 system.3. Select Standard (~ 2 hours to complete a run) for the ramp speed.4. Select cDNA (complementary DNA) for the template type. (default)5. Click Next.1243
