Prepare The Reaction Mix - abi 7500 Getting Started Manual

Chapter 7 Prepare the Comparative C

Prepare the Reaction Mix

Prepare the Reaction Mix

About the
Example
Experiment
Required
Materials
Notes
144
Applied Biosystems 7500/7500 Fast Real-Time PCR System Getting Started Guide for Relative Standard Curve
Reactions
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Prepare the reaction mix using the components and volumes that were calculated by the
7500 software ("Complete the Reaction Mix Calculations Tab for the Assays" on page 127).
The software calculates the volumes for all components for the PCR reactions.
Note:
However, when you prepare the reaction mix in this section, include only the master mix,
assay mix, and water. Add the sample when you prepare the reaction plate (see "Prepare
the Reaction Plate" on page 146).
For the comparative C example experiment:
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• The reaction mix components are:
®
– TaqMan Universal PCR Master Mix (2✕)
– 18S Assay Mix (20✕)
– EGR3 Assay Mix (20✕)
– MAOB Assay Mix (20✕)
– OGDH Assay Mix (20✕)
– OSGEP Assay Mix (20✕)
– SERPING1 Assay Mix (20✕)
– Water
• The volumes calculated in the software for the target assays are:
Component
Master Mix (2.0✕)
Assay Mix (20.0✕)
Water
Total Volume
The sample is not added at this time.
Note:
• Microcentrifuge tubes
• Pipettors
• Pipette tips
• Reaction mix components (listed above)
• Centrifuge
Volume (µL) for 1 Reaction
25.0
2.50
17.50
45.0
and Comparative C Experiments
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