Increment/Decrement Time; Temperature Gradient; Graphical Representation - BIO RAD MyCycler Instruction Manual

4.3.2.2 Increment/Decrement Time
Most protocols call for the exact same dwell time for each repeat of a cycle, but
in some instances you may wish to program an automatic increase or decrease in
the step dwell time after each repeat. To input an automatic increment or decrement
function for a step, the cycle containing the step must have more than one cycle
repeat. The automatic increment and decrement function updates the dwell time by
a specified amount, after every repeat, starting after repeat one.
For example, you could create a cycle with 3 repeats, with a first step at 95°C
for 10 seconds and a second step at 55°C for 45 seconds. You could then specify
a 5-second automatic dwell time decrement for the second step. Thus, on the first
repeat of the cycle, the instrument would hold at 55°C for 45 seconds. Then on the
second repeat of the cycle, the instrument would hold at 55°C for 40 seconds.
Finally, on the third repeat of the cycle, the instrument would hold at 55°C for 35
seconds.
Dwell time increments or decrements may be as low as 1.0 seconds, and they
may be as long as desired, so long as they do not result in values outside the legal
ranges described in section 4.3.1. If you input an increment or decrement function
that would result in an illegal dwell time value, the thermal protocol will run, but the
dwell time will cease to increment or decrement once the upper or lower limit has
been reached. Furthermore, for every repeat that would result in an illegal dwell
time value, an error is counted on the End of Run screen and displayed in the
Validation and Run reports.
4.3.2.3 Temperature Gradient
The MyCycler Thermal Cycler offers a temperature gradient option for
optimization of assay conditions. Gradients may be programmed at any step of any
cycle of the protocol. The gradient option allows input of a temperature gradient of
1-25 degrees in total temperature span at temperatures as low as 30°C and as
high as 99°C.
Thermal Gradients in the MyCycler are formed from the front to the rear of the
sample block resulting in eight specific temperatures for assay optimization. This
innovative design allows for evaluation of the maximum assay components at each
specific temperature. Each row includes twelve wells for evaluation of assay
components such as MgCI2 or primer concentrations.
The temperature gradients formed by the MyCycler take a non-linear yet highly
reproducible shape that allows for excellent prediction of actual sample temperatures
at each row. Extensive validation of the gradient temperatures indicate that the non
linear gradient is both predictable and reproducible.
4.4 Graphical Representation
Thermal protocols are displayed graphically as a temperature plot. The graphical
representation shows a left-to-right plot of the thermal cycling program and reflects
approximate values of the set point temperatures along its y-axis. The plot displays
the set point temperature, dwell times, and advanced options, such as time or
temperature increments and decrements, for each step.
At each step, the set point temperature in Celsius (e.g., 95.0°C) is shown immediately
above the temperature plot, and the dwell time (in mm:ss format) is shown immediately
below the temperature plot. Temperature increments and decrements are denoted within
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