For both the Pacific and Atlantic samples, the log2 RT-qPCR normalized value and the log2 microarray value for each sample were correlated in R (R Core Team 2014) using linear models with microarray values as the independent variable and RT-qPCR values as the dependent variable to obtain an adjusted R-squared value and slope of the relationship for each gene.
This was done by comparing the correlation metric to the mean absolute expression level and the standard deviation, both for microarray values and real-time PCR values.
While c-fos and Egr1 expression at 3 mg/kg deltamethrin and 100 mg/kg permethrin reflect the direction of fold-change observed in the microarray study, very little similarity is apparent between qRT-PCR and microarray expression values for these genes at the lower dose levels (Table 4).
Robust Microarray Analysis (RMA) background correction and quantile normalization were used to generate microarray expression values for the Affymetrix platform data sets, EIF4G1, RhoA, and CTNNB1.
For the RNA-Seq assembled transcripts (each transcript was regarded as a gene model, although it may be only a transcription fragment), all previous designed microarray probes (10) were re-mapped to the RNA-Seq assembled transcripts, and the microarray expression values for these transcripts were re-normalized as in the study by Miao et al. (10) and provided in the TetraFGD.
The results (FC, p-value and FDR) of the performed comparative analyses for all the probe sets encoding AMP genes are given in Table S2. Figure 1A F shows the individual microarray expression values for DEFB1, LEAP2, PYY, DEFA5, DEFA6 and DEFB4, respectively.
Microarray RMA values for each transcript cluster for each of 9 samples were submitted to a Principal Component Analysis in order to detect possible outliers.
Microarray expression values for Medicago and Lotus were downloaded from http://mtgea.noble.org/v3/ [ 49, 50] and http://ljgea.noble.org/ v2/ [ 51].
Microarray expression values for potential biomarker genes were regressed against corresponding liver Se concentrations using the Excel 2007 Data Analysis add-in, as described previously [ 30].
Raw and normalized microarray expression values for each probe were plotted on a log-scale histogram to represent the distribution of mRNA expression in the undifferentiated mESC transcriptome.
