Eppendorf Mastercycler Gradient - Programming Examples Overview; Gradient PCR Example

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7 Programming
7.6 Examples of programming
The following passage contains several examples of the versatility of the commands and functions of the
Mastercyclers.
7.6.1 Gradient PCR (for Mastercycler
®
gradient only)
The Gradient PCR is used to optimize temperatures in a PCR experiment. The gradient may be programmed
with a temperature range of up to 20 °C with every temperature command.
The most common application is the determination of the optimal annealing temperature (see example), for
which a gradient of, for example, ± 10 °C is built up. The recommended mean gradient is 5 °C higher than the
calculated annealing temperature.
Gradient PCR
Temperature control of tubes Control tube
Temperature control of lid lid 105°
Nowait auto
1 Initial denaturation 94° 2 min
2 Denaturation 94° 15 sec
3 Annealing 60° 15 sec
Gradient G=10 °
4 Elongation 72° 30 sec
5 Number of cycles go to 2 Rep 29
6 Final elongation 72° 2 min
7 Cooling and storage Hold 20° enter
End end