Fig. 2: RNA purification methods strongly influence mRNA and miRNA sequencing.

Performance metrics are shown for both mRNA capture (left panels) and miRNA (right panels) sequencing. For each unique RNA purification-plasma input volume combination, 3 technical replicates were analyzed (n = 39 for mRNA capture & n = 45 for miRNA sequencing). Absolute numbers of detected mRNAs (a) and miRNAs (b) that reached the count threshold (see “Methods”) are shown. High numbers indicate good performance. Endogenous mRNA (c) and miRNA (d) concentration. Values are log rescaled to the lowest mean of all methods and transformed back to linear scale. The mean and 95% confidence interval are shown. High concentrations indicate good performance. Replicate variability based on ALC at mRNA (e) and miRNA (f) level, respectively. Small ALC indicates good performance. Overview of all performance metrics at mRNA capture (g) and miRNA (h) sequencing level, respectively, after transforming the values to robust z-scores. High z-scores indicate good performance. Rows and columns of the heatmaps are clustered according to complete hierarchical clustering based on Euclidean distance. Average z refers to the mean of robust z-scores for a specific RNA purification method. The number that follows the name of the purification method is the plasma input volume (in ml). MAX=Maxwell method, MIR=miRNeasy method, MIRA=miRNeasy Advanced method, MIRV=mirVana method, MIRVE=mirVana method with purification protocol for small RNA enrichment, NOR=Norgen method, NUC=NucleoSpin method, QIA=QIAamp method.