Operator`s manual

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LightCycler® 480 Gene Scanning Software
Preparing a LightCycler® 480 System Gene Scanning Experiment
Rapid Detection of Amplification Artifacts
Rapid Detection of Amplification Artifacts6
The goal of a LightCycler® 480 System Gene Scanning assay is to generate a single pure
PCR product. Any amplification artifacts (e.g., primer-dimers) may lead to misleading
results.
On the LightCycler® 480 System, you can easily determine whether the PCR products
include primer-dimers by comparing your samples to a non-template control. After the
initial melting curve run, simply view the Melting Peaks plot (negative first derivative of
the sample fluorescence plotted against temperature; generated by the T
m
Calling Analysis
Module of the LightCycler® 480 Software). That plot should show no peaks for the no-
template control. If initial experiments show reaction product in your no-template con-
trol, reoptimize the reaction (e.g., by designing new primers) and repeat the experiment.
For further verification, you may also want to examine the product on an electrophoretic
gel.
You can access the Melting Peaks plot directly from the LightCycler® 480 Gene Scanning
Software module. There is no need to run a separate T
m
Calling Analysis.