The three samples from each patient were compared with 11 in vitro microarray time points with highest correlation [ 4].
From each cultivation experiment, biomass was harvested from the mid-exponential growth phase (biomass concentration 1.5 gr/kg) and used for RNA extraction and subsequent microarray analysis (time point 1; HacAWT-1; [glucose] = 5.8 g/L).
The Sample Information File was created, which, besides the creation date, contains the microarray name, time point (0, 1, 2, 4, and 12 h), and treatment (IL-1 β, TNF- α, TGF- β, and PDGF-D) of each sample, all serving as covariates for the ComBat method.
After quality analysis and normalization (described above), we had a three-point LD time course with four microarrays per time point (2 independent biological replicates and 2 dye-swap experiments).
To ensure that the RNA-Seq data could be compared with the microarray data, time points were selected a subset of those in the previous microarray expression studies that covered 20 states of the three stages of the T. thermophila life cycle.
Microarray samples were available for 9 time points (0, 3, 6, 12, 24, 48, 72, 120 and 192) h after stimulation, with 3 (mRNA microarray) and 2 (miRNA microarray) replicates per time point.
We reanalyzed their microarray data (four Arabidopsis ATH1 microarrays, two time points, no biological duplication) with the same data processing methods (1.2-fold cutoff threshold) used in this study.
For microarray studies, each time point was represented by three biological replicates analysed in a dye swap design (six hybridizations per time point).
After E. coli challenge, the microarray results at time point 24 h were in agreement to the respective results obtained with RT PCR [ 12].
As in the microarray analyses, a time point close to 2 weeks was chosen in order to maximise the effects of Nox4 signaling apparent in the hearts.
