DNA Engine Tetrad 2 Thermal Cycler Operations Manual
5-24
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Step two of our protocol should now specify a temperature incubation of 92.0 for a
duration of 00:30.
Entering a Gradient Step
Step three of our program is a gradient step designed to determine the optimal
annealing temperature of our oligonucleotide. If step three of the displayed protocol
is not a gradient step (two black lines):
•
Select step two.
•
Select <Grad>.
A new gradient step should now appear as step three with the default higher temperature
limit of 65°C and the default lower temperature limit of 50°C. The default duration is 30
seconds.
The maximum temperature range for a gradient is 24°C and the minimum is 1°C.
Fractional degrees are not accepted.
Recall that in our example protocol, the gradient step should specify a range of 45°C
to 65°C and a duration of 3 minutes.
•
Position the cursor (text pointer) in the higher limit temperature field and
select by clicking once.
•
Use the backspace key on the numeric keypad to delete the higher tem-
perature if it is not 65°C.
•
Enter 65 from the numeric keypad.
•
Repeat the steps above for the lower temperature limit, entering a value
of 45 from the keypad.
•
Click once to accept the changes to the gradient range.
Note:
Change both the higher and lower temperatures before accepting the
changes to the gradient step to ensure that the temperature differential is not
greater than 24°C or less than 1°C.
•
Position the cursor (text pointer) in the time:minute field of step 3 and
select by clicking once.
Step 2
temperature step
