Rotor-Gene Q MDx CE User Manual 02/2022
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To set up a Melt, specify the start temperature, the end temperature, the temperature increments,
the length of time to hold at the first acquisition temperature before the ramp is initiated, the time
each increment is to be held for, and the acquisition channels.
A ramp will be generated between the 2 temperatures. If the start temperature is higher than the
end temperature, the name of the step will change to
Hybridisation
. The
Acquiring To
option, set to
Melt A in the screenshot below, can be changed by clicking the button. The
Acquisition
window
will appear and the channels can be selected.
When running a standard melt the temperature is increased by increments of 1°C, waiting for
5 seconds before each acquisition. The Rotor-Gene Q MDx can be configured to perform melts in
0.02°C increments. The minimum hold time between temperature steps varies depending on the
number of degrees between each step.
High Resolution Melt
High resolution melt (HRM) analysis characterizes double-stranded DNA samples based on their
dissociation (melting) behavior. It is similar to classical melting curve analysis, but provides far more
information for a wider range of applications. Samples can be discriminated according to
sequence, length, GC content, or strand complementarity, down to single base-pair changes.
HRM analysis can only be performed on instruments that have HRM hardware and software
installed. Data is acquired using specialized HRM sources and detectors. HRM analysis also
includes the option to perform Gain Optimisation just before the Melt begins. After performing
HRM, the data can be analyzed with HRM analysis software (Section 10).
Optical Denature Cycling
Optical Denature Cycling is an exciting technique, available on the Rotor-Gene Q MDx, which
performs real-time melt analysis to determine the melt peak of a reference sample. This indicates
PCR product denaturation with higher precision than setting a particular denature temperature for
a hold time. To perform this technique, simply place a reference tube of PCR product in tube position
1 of the rotor. The reference tube must also contain a detection chemistry that enables detection of
strand dissociation.