User manual

Droplet Digital
™
PCR Applications Guide | 25
Designing Droplet Digital
™
PCR Experiments
In Figure 2.13, a gradient of annealing/extension temperatures is shown for three amplicons
with different lengths: A, B, and C correlate to 200, 99, and 62 bp amplicons, respectively.
The experiment demonstrates the effect amplicon length has on fluorescence amplitude
as well as how annealing/extension temperature affects amplitude and the ability to resolve
products with different lengths based on amplicon size. Red arrows show increasing
separation between 200 and 99 bp amplicons as the annealing/extension temperature
is lowered from 65 to 55˚C.
0 100,000 200,000 300,000
Event number
Channel 1 amplitude
40,000
35,000
30,000
25,000
20,000
15,000
10,000
5,000
0
A04
A B C A B C A B C A B C A B C A B C A B C A B C
A05 A06 B04 C04 D04 E04 F04 G04 H04B05 C05 D05 E05 F05 G05 H05B06 C06 D06 E06 F06 G06 H06
65°C 55°C
Fig. 2.13. Gradient of annealing/extension temperatures for three amplicons with different lengths:
A (200 bp), B (99 bp), and C (62 bp).