abi 7500 - Prepare the Reaction Plate

Chapter 7 Prepare the Comparative C
T
Reactions
Prepare the Reaction Plate
Applied Biosystems 7500/7500 Fast Real-Time PCR System Getting Started Guide for Relative Standard Curve
and Comparative C
T
Experiments
146
Notes
For More
Information
For more information on preparing the reaction mix, refer to the protocol appropriate for
the reagents you are using in the PCR reactions:
•TaqMan
®
Gene Expression Assays Protocol
• Custom TaqMan
®
Gene Expression Assays Protocol
Prepare the Reaction Plate
Prepare the reactions for each replicate group, then transfer them to the reaction plate.
Use the plate layout displayed in the 7500 software.
About the
Example
Experiment
For the comparative C
T
example experiment:
•A MicroAmp
®
Optical 96-Well Reaction Plate is used.
• The reaction volume is 50 µL/well.
• The reaction plate contains:
– 96 Unknown wells
– 0 Negative Control wells
– 0 Empty wells
• The plate layout that is automatically generated by the 7500 software is used:
Required
Materials
• Microcentrifuge tubes
• Pipettors
• Pipette tips
• Reaction mixes (from page 145)
•Water
• Samples (from page 143)
•MicroAmp
®
Optical 96-Well Reaction Plate
•MicroAmp
™
Optical Adhesive Film
• Centrifuge