Prepare The Standard Dilution Series - abi 7500 Getting Started Manual

Prepare the Standard Dilution Series

You prepare the standard dilution series using the volumes that were calculated by the 7500
software ("To complete the Reaction Mix Calculations tab for the c-myc assay" on page 37
and "To complete the Reaction Mix Calculations tab for the GAPDH assay" on page 38):
About the
For the relative standard curve example experiment:
Example
• The standard (stock) concentration is 100 ng/µL.
Experiment
• The volumes calculated in the software for both the c-myc and GAPDH assays are:
Dilution Point
Source
(Factor)
1 (200) Stock
2 (20) Dilution 1
3 (2) Dilution 2
4 (0.2) Dilution 3
5 (0.02) Dilution 4
Required
• Water (to dilute the standards)
Materials
• Microcentrifuge tubes
• Pipettors
• Pipette tips
• Standard stock
• Vortexer
• Centrifuge
Prepare the 1. Label a separate microcentrifuge tube for each standard:
Standard Dilution
Series for the
c-myc Assay
Notes
Applied Biosystems 7500/7500 Fast Real-Time PCR System Getting Started Guide for Relative Standard Curve
and Comparative C Experiments
T
Chapter 3 Prepare the Relative Standard Curve Reactions
Source Volume Diluent Volume
(µL) (µL)
8.07 12.10
2.02 18.15
2.02 18.15
2.02 18.15
2.02 18.15
• c-myc Std. 1
• c-myc Std. 2
• c-myc Std. 3
• c-myc Std. 4
• c-myc Std. 5

Prepare the Standard Dilution Series

Total Volume Standard Concentration
(µL) (ng/µL)
20.17 40.0
20.17 4.0
20.17 0.4
20.17 0.04
20.17 0.004
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