Sample sets and Ranking of candidate reference genes as calculated by geNorm. Left panel: Independent cultures (illustrated by the boxes) were carried out: Wild type KT strain on glucose (sample set B) and galactose (set C); Wild type CEN.PK (set D & set E as independent cultures) and its tps1 derivative strain (set H & set I as independent cultures), on galactose. Sampling [S#] was performed all along the cultures with a posteriori selection and analysis of 4 to 7 RNA samples representative of different physiological states (e.g. samples 1 to 5 for growth of the KT strain on glucose; see Additional file 1). Expression data from one culture (e.g. set B/) or from several cultures (connector between boxes, e.g. set A/that includes samples from sets B & C together) were then analyzed with geNorm (A ~M sample sets). Right panel: Synthetic overview of ranking of the candidate reference genes according to their expression stability, and determination of the optimal number of genes used for normalization. The 2 most stable genes (black circle), the third (dashed circle) and the following 3 best reference genes (empty circle). Pair-wise variation (Vn/n+1) between NFn and NFn+1 (NF: normalization factor; n: number of genes used for NF calculation). Right Column: pair-wise variation value below the threshold 0.15, which means that n genes might be sufficient for NF calculation (i.e. 2 genes for set "A"). See additional file 3 for overall stability under the standard geNorm output format.