Skip to main content

Table 1 Transgene copy number determination by different methods.

From: Statistical tools for transgene copy number estimation based on real-time PCR

Line Zygosity Method LgRt(T/C) Copy Num Confidence Interval Efficiency Adjust Pred Copy Exp Copy
1 Hemi-zygous 1 -1.05 0.49 (0.42, 0.56) N/A 0.5 0.5
   2 -1.07 0.48 (0.33, 0.68) No 0.5 0.5
   3 -0.93 0.53 (0.32, 0.42) Yes (0.93) 0.5 0.5
   4 -1.00 0.5 (0.12, 2.23) N/A N/A 0.5
  Homo-zygous 1 -0.10 0.93 (0.79, 1.09) N/A 1 1
   2 -0.06 0.96 (0.77, 1.19) No 1 1
   3 0.01 1.01 (0.85, 1.21) No 1 1
   4 -0.80 0.57 (0.24, 1.38) N/A N/A 1
2 Hemi-zygous 1 -1.10 0.47 (0.41, 0.52) N/A 0.5 0.5
   2 -1.18 0.44 (0.21, 0.95) No 0.5 0.5
   3 -1.25 0.42 (0.33, 0.52) No 0.5 0.5
   4 0.06 1.04 (0.22, 4.99) N/A N/A 1
  Homo-zygous 1 0.19 1.14 (0.93, 1.40) N/A 1 1
   2 -0.03 0.98 (0.81,1.19) No 1 1
   3 0.18 1.13 (0.88, 1.47) Yes (0.97) 1 1
   4 0.09 1.06 (0.31, 3.68) N/A N/A 1
  1. The transgene copy numbers for both homozygous and hemizygous progenies of two transgenic lines were analyzed with real-time PCR and different statistical methods. Method 1 is the external standard curve design, Method 2 is the internal reference gene design with standard curve, Methods 3 and 4 are the internal reference gene design with fluorescence data. In Method 3, the cycle number was used as dependent variable, and in Method 4, the logarithm-transformed fluorescence signal was used as the dependent variable. Column 4 shows the logarithm-2 transformed ratio between the test line and control line, or between the transgene and internal reference gene. Column 4 presents the direct outputs from SAS code. Column 5 indicates the numeric estimation of copy number. The confidence levels for logarithm 2 transformed ratios are converted into confidence interval for transgene copy number and presented in Column 6. Column 7 indicates whether efficiency adjustment was included or not. The predicted and actual copy numbers were presented in Column 8 and 9.