Four PCR cycles apart means 16 times the DNA
- The idea
- Predict
- See it
- Why
- Check
The idea
“A sample that crosses the threshold 4 cycles sooner started with about 4 times the DNA.”
It's easy to count cycles like steps on a ruler, each one adding the same amount. In PCR, each cycle doubles what's already there.
The short version
The mistake: A sample that crosses the threshold 4 cycles sooner started with about 4 times the DNA.
What's actually true: Each cycle doubles the DNA, so a gap of n cycles means 2^n times the starting DNA. Four cycles is 2 × 2 × 2 × 2 = 16 times.
- Each cycle heats the DNA to separate its strands. A heat-stable DNA polymerase copies each strand from primers. Each copy becomes a template, so copies ideally double.
- Twice the starting DNA puts a sample one cycle ahead. Four times puts it 2 cycles ahead, and 16 times puts it 4 ahead.
- In real-time PCR, a dye glows brighter as copies build up. The threshold cycle is when the glow first passes a set level.
- Late in a run, the curves flatten as primers and nucleotides run low. Every curve here has the same shape, just shifted, so the gap is the same at any height.
On the exam: Cycle gaps are doublings. Samples n cycles apart differ 2^n-fold, and the one that crosses first had more.