Real-Time Quantitative RT-PCR: Design, Calculations, and Statistics

Two recent letters to the editor of The Plant Cell ([Gutierrez et al., 2008][1]; [Udvardi et al., 2008][2]) highlighted the importance of following correct experimental protocol in quantitative RT-PCR (qRT-PCR). In these letters, the authors outlined measures to allow precise estimation of gene

[1]  Christian Ritz,et al.  Highly accurate sigmoidal fitting of real-time PCR data by introducing a parameter for asymmetry , 2008, BMC Bioinformatics.

[2]  L. R. Verdooren Statistical procedures for agricultural research: (2nd edition) K.A. Gomez and A.A. Gomez. John Wiley, New York, 1984. xvi + 680 pp., US$39.95. John Wiley, Chichester, Sussex, U.K., £37.95. Also paperback £12.95/US$15.50. ISBN 0-471-87092-7 , 1986 .

[3]  O. Van Wuytswinkel,et al.  Towards a Systematic Validation of References in Real-Time RT-PCR , 2008, The Plant Cell Online.

[4]  D. F. Cox,et al.  Statistical Procedures for Agricultural Research. , 1984 .

[5]  Mariano J. Alvarez,et al.  Model based analysis of real-time PCR data from DNA binding dye protocols , 2007, BMC Bioinformatics.

[6]  G. Mortier,et al.  qBase relative quantification framework and software for management and automated analysis of real-time quantitative PCR data , 2007, Genome Biology.

[7]  R. Mead,et al.  The Design of Experiments: Statistical Principles for Practical Applications. , 1989 .

[8]  The Design of Experiments: Statistical Principles for Practical Application. , 1989 .

[9]  K. Voelkerding,et al.  Evaluation of quantification methods for real-time PCR minor groove binding hybridization probe assays. , 2007, Analytical biochemistry.

[10]  Jonathan C. McDowell,et al.  Data analysis software , 1997, Current Opinion in Neurobiology.

[11]  A. Moorman,et al.  Assumption-free analysis of quantitative real-time polymerase chain reaction (PCR) data , 2003, Neuroscience Letters.

[12]  H. D. Patterson,et al.  Recovery of inter-block information when block sizes are unequal , 1971 .

[13]  W. Scheible,et al.  Eleven Golden Rules of Quantitative RT-PCR , 2008, The Plant Cell Online.

[14]  A. Moorman,et al.  Amplification efficiency: linking baseline and bias in the analysis of quantitative PCR data , 2009, Nucleic acids research.

[15]  Eun-Soo Han,et al.  SAS programs for real-time RT-PCR having multiple independent samples. , 2004, BioTechniques.