Relative transcript quantification by Quantitative PCR: Roughly right or precisely wrong?
Journal article, Peer reviewed
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Original versionBMC Molecular Biology 2005, 6:10, 1-3 10.1186/1471-2199-6-10
Background When estimating relative transcript abundances by quantitative real-time PCR (Q-PCR) we found that the results can vary dramatically depending on the method chosen for data analysis. Results Analyses of Q-PCR results from a salmon louse starvation experiment show that, even with apparently good raw data, different analytical approaches [1, 2] may lead to opposing biological conclusions. Conclusion The results emphasise the importance of being cautious when analysing Q-PCR data and indicate that uncritical routine application of an analytical method will eventually result in incorrect conclusions. We do not know the extent of, or have a universal solution to this problem. However, we strongly recommend caution when analysing Q-PCR results e.g. by using two or more analytical approaches to validate conclusions. In our view a common effort should be made to standardise methods for analysis and validation of Q-PCR results.