In order to get an overview of the reliability of the microarray data we performed RT-PCRs of 17 genes.
Therefore, qRT-PCR was performed for 16 ABC transporter genes to evaluate the reliability of the microarray data (Table 3).
The results from microarray analysis and qRT-PCR showed the same patterns for either differentially increased or decreased expression of the genes, suggesting the reliability of the microarray analysis.
Correlation coefficient (r) values were 0.662, 0.728, and 0.678 at the P ≤ 0.04 level for the WTd-WTw, T2d-T2w, and T2d-WTd experiments, respectively, indicating the reliability of the microarray data.
To confirm the reliability of the microarray data and the robustness of the strategy for identifying genes with altered expression, we examined the protein levels of the identified genes using immunohistochemical analysis of human tissues.
To determine the reliability of the microarray data from the chloroplast genes, a two-step qRT-PCR was performed on the RNA material used in the microarray analyses for time points 0.5 h, 3 h, 12 h and 24 h.
The validation assay showed strong correlation of gene expression with the microarray results under all tested conditions (r>0.9, Figure S2), confirming independently the reliability of the microarray data.
The reliability of the microarray expression data was assessed by quantitative real-time reverse transcriptase PCR (qRT-PCR) analysis.
To determine the reliability of the microarray results, we analyzed the expression of 13 selected genes from the MHC and NKC regions by qRT-PCR experiments in 8 of the sample pairs that had been used for the microarrays (see Fig. 1).
These results confirmed the reliability of the microarray data.
