How many biological replicates do I need for my RNA-Seq experiment?

rna-seqIn the design of microarray or next-generation sequencing experiments it is crucial to choose the appropriate number of biological replicates. As often the number of differentially expressed genes and their effect sizes are small and too few replicates will lead to insufficient power to detect these. On the other hand, too many replicates unnecessary leads to high experimental costs. Power and sample size analysis can guide experimentalist in choosing the appropriate number of biological replicates. Several methods for power and sample size analysis have recently been proposed for microarray data. However, most of these are restricted to two group comparisons and require user-defined effect sizes.

Here a team led by researchers at the Leiden University Medical Center, The Netherlands propose a pilot-data based method for power and sample size analysis which can handle more general experimental designs and uses pilot-data to obtain estimates of the effect sizes. The method can also handle χ2 distributed test statistics which enables power and sample size calculations for a much wider class of models, including high-dimensional generalized linear models which are used, e.g., for RNA-seq data analysis. The performance of the method is evaluated using simulated and experimental data from several microarray and next-generation sequencing experiments. Furthermore, the researchers compare their proposed method for estimation of the density of effect sizes from pilot data with a recent proposed method specific for two group comparisons.

Availability – SSPA is available at:

  • van Iterson M, van de Wiel MA, Boer JM, de Menezes RX. (2013) General power and sample size calculations for high-dimensional genomic data. Stat Appl Genet Mol Biol 12(4)4, 49-67. [abstract]