A simulation approach to assess the minimal number of real-time PCR replicates for GM quantification

The quantification of genetically modified (GM) ingredients in food and feed typically uses real-time quantitative PCR (RT-QPCR). In recent years a multitude of new RT-QPCR assays have facilitated increased method performance. The level of sample replication within these assays is a fundamental aspect that needs to be considered to produce results with high confidence. In this paper we describe the use of a modelling approach as applied to GM and RT-QPCR data sets, to objectively assesses the effect of different levels of PCR replication in terms of the variability associated with a result, and demonstrate that it is possible to use a reduced level of replication without a subsequent reduction in the confidence of a result. Using an example data set, we show it is possible to reduce the sample level of replication from six to three PCR replicates, without a significant change in the mean value of the result. The use of such an approach can facilitate the use of the minimum number of replicates in order to produce an accurate result, thus saving on important resources involved in quantification assays.