• Dilute the dye in buffer compatible with your Master Mix.
•
(Intercalating dyes only)
Add the appropriate amount of amplified PCR product to
generate fluorescence.
Prepare a custom dye dilution plate
IMPORTANT!
Wear powder-free gloves throughout the procedure.
1.
Prepare a 2- or 3-fold dilution series of the custom dye.
2.
Dispense aliquots of each dilution into the center of a reaction plate, then seal
the plate.
A full plate is not needed.
100 nM
200 nM
400 nM
800 nM
1600
nM
100
nM
200
nM
400
nM
800
nM
1600 nM
100
nM
200
nM
400
nM
800
nM
1600 nM
3.
Vortex the plate for 5 seconds, then centrifuge at 750–1,000 ×
g
for 2 minutes.
4.
Confirm that the liquid in each well is at the bottom of the well and free of
bubbles. If it is not, centrifuge the plate again.
Chapter 7
Calibrate and verify instrument performance
Calibrate custom dyes
7
QuantStudio
™
6 Pro and 7 Pro Real-Time PCR Systems User Guide
99
