Quantitative
"Real Time"
RT-PCR as a " + / - "
test
Progress of DNA amplification during a Polymerase
Chain Reaction (PCR)
can be
monitored in "real
time" (RT-PCR) by
measuring the release of fluorescent "flashes" during
amplification. Reaction rates can be measured continuously, or
determined at a fixed time-point during the
exponential amplification phase. A computer measures the rate of
"flashing" in 96 simultaneous experimental PCR reactions relative to a control reaction
in
the same tube (top, right). A
"positive"
result ("red"
signals, below right) indicates that the
target DNA sequence is
present. Because amplification is measured against a control, a "negative"
("black" background
signal) is a
definite indication of the absence of the
target allele.
Unlike ordinary preparative PCR, RT-PCR
allows the success of multiple PCR
reaction to be determined
automatically after only
a few cycles, without separate analysis of each reaction, and
avoids
the
problem of "false negatives".
