Operator`s manual

14
LightCycler® 480 Gene Scanning Software
Gene Scanning with the LightCycler® 480 System
Gene Scanning with the A
LightCycler
®
480 System
“Gene scanning” or “mutation scanning” techniques detect the presence of sequence
variation in target-gene derived PCR amplicons. “Gene Scanning” is based on “High
Res olution Melting”, a novel, closed-tube post-PCR (Polymerase Chain Reaction) method
enabling genomic researchers to analyze genetic variations in PCR amplicons prior to
or as an alternative to sequencing. High Resolution Melting provides high specificity,
sensi tivity and convenience at significantly higher speed and much lower cost than other
established (e.g., gel-based) methods. For example, in a diploid genome, equivalent re-
gions from maternal and paternal chromosomes are both amplified by the Polymerase
Chain Reaction. The PCR products can then be analyzed for completely matched hybrids
(called homoduplexes) and mismatched hybrids (heteroduplexes).
In a LightCycler® 480 System Gene Scanning experiment, sample DNA is first ampli-
fied via real-time PCR in the presence of LightCycler® 480 High Resolution Melting
Dye. Immediately after DNA amplification, a High Resolution Melting experiment can
be per formed on the same LightCycler® 480 Instrument and analyzed with LightCycler®
480 Gene Scanning Software to identify sequence variants. Thus, the entire mutation
screening process is homogeneous. That is, the entire experiment can be done on the
LightCycler® 480 Instrument; post-PCR analysis does not require a separate device.