RT-qPCR study on post-mortem brain samples from patients with major psychiatric disorders: Reference genes and specimen characteristics

BackgroundGene expression studies conducted in post-mortem human brain samples have the potential to identify relevant genes implicated in psychiatric disorders. Although reverse transcription quantitative real-time PCR (RT-qPCR) has emerged as the method of choice for specific gene expression studies, it requires the use of stable reference genes, and it is necessary to control for pre- and post-mortem factors to obtain reliable data.ObjectiveThe aim of this study was to identify suitable reference genes and specimen characteristics that can be taken into account when comparing mRNA expression data between post-mortem brain specimens from psychiatric patients and controls.MethodWe used a selection of suitably matched occipital cortex specimens from subjects in each of the following groups: schizophrenia (N = 15), bipolar disorder (N = 13), major depressive disorder (N = 15), and control (N = 15). Quantitative and qualitative RNA analyses were performed prior to RT-qPCR and gene expression stability was evaluated with geNorm and NormFinder.ResultsWe identified GAPDH, RPS17, RPL30, RPLP0, and TFRC as potential reference genes from a sample plate containing 32 candidates commonly used as reference genes. Further analyses of these 5 genes highlighted that 1) they are suitable reference genes for RT-qPCR studies in these post-mortem brain samples from psychiatric patients, and 2) the RNA quality index is highly correlated with gene expression values (r = −0.681, p < 0.0001).ConclusionsIn addition to controlling for pre- and post-mortem factors and selecting stable reference genes for normalization, sample sets should be matched with regard to RNA quality.