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Table 1 An example of four-parameter sigmoidal equation modeling and result of fluorescence normalization

From: Quantify single nucleotide polymorphism (SNP) ratio in pooled DNA based on normalized fluorescence real-time PCR

Cycle Numberxa D2 sample FAM (Raw Rx)b D2 Normalized Rx . (Raw Rx -/Raw Y0-1)c
23.58 353.48 0.056045583
24.58 355.38 0.061721963
25.58 f 367.68 0.098469051
26.58 380.74 0.13748669
27.58 409.2 0.222512879
28.56 429.79 0.284026907
29.61 465.57 0.390922095
30.56 504.15 0.506182473
31.63 535.18 0.598886713
32.63 576.31 0.72176539
33.63 595.56 0.779276076
34.63 639.21 0.909683425
35.63 669.3 0.99957935
Parameter Sigmoid function fitted for raw fluorescenced Sigmoid function fitted for normalized fluorescencee
a 525.5884 1.5702
b 3.2674 3.2674
X0 33.4481 33.4481
Y0 334.7204 0
R 0.998805 0.99880514
  1. a amplification cycle number
  2. b Raw FAM fluorescence data
  3. cNormalized fluorescence = Raw Rx/Raw yo-1
  4. d 4 parameter sigmoid function fitted for raw fluorescence
  5. e 4 parameter sigmoid function fitted for normalized fluorescence
  6. f bold text cycles are selected PCR exponential phase for allele ratio quantification