Evaluation of gene expression levels by reverse transcription quantitative real-time PCR (RT-qPCR) has for many years been the favourite approach for discovering disease-associated alterations. Normalization of results to stably expressed reference genes (RGs) is pivotal to obtain reliable results. This is especially important in relation to neurodegenerative diseases where disease-related structural changes may affect the most commonly used RGs. We analysed 15 candidate RGs in 98 brain samples from two brain regions from Alzheimer’s disease (AD), Parkinson’s disease (PD), Multiple System Atrophy, and Progressive Supranuclear Palsy patients. Using RefFinder, a web-based tool for evaluating RG stability, we identified the most stable RGs to be UBE2D2, CYC1, and RPL13 which we recommend for future RT-qPCR studies on human brain tissue from these patients. None of the investigated genes were affected by experimental variables such as RIN, PMI, or age. Findings were further validated by expression analyses of a target gene GSK3B, known to be affected by AD and PD. We obtained high variations in GSK3B levels when contrasting the results using different sets of common RG underlining the importance of a priori validation of RGs for RT-qPCR studies.
Electronic supplementary material
The online version of this article (doi:10.1038/srep37116) contains supplementary material, which is available to authorized users.
About this article
Cite this article
Rydbirk, R., Folke, J., Winge, K. et al. Assessment of brain reference genes for RT-qPCR studies in neurodegenerative diseases. Sci Rep 6, 37116 (2016). https://doi.org/10.1038/srep37116
Journal of Biotechnology (2021)
Frontiers in Molecular Neuroscience (2021)
Assessment of common housekeeping genes as reference for gene expression studies using RT-qPCR in mouse choroid plexus
Scientific Reports (2021)
Increased transcription of TSPO , HDAC2 , and HDAC6 in the amygdala of males with alcohol use disorder
Brain and Behavior (2021)
GABAergic neurons are susceptible to BAX-dependent apoptosis following isoflurane exposure in the neonatal period
PLOS ONE (2021)