Assessment of brain reference genes for RT-qPCR studies in neurodegenerative diseases
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Assessment of brain reference genes for RT-qPCR studies in neurodegenerative diseases. / Rydbirk, Rasmus; Folke, Jonas; Winge, Kristian; Aznar, Susana; Pakkenberg, Bente; Brudek, Tomasz.
In: Scientific Reports, Vol. 6, 37116, 17.11.2016.Research output: Contribution to journal › Journal article › Research › peer-review
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T1 - Assessment of brain reference genes for RT-qPCR studies in neurodegenerative diseases
AU - Rydbirk, Rasmus
AU - Folke, Jonas
AU - Winge, Kristian
AU - Aznar, Susana
AU - Pakkenberg, Bente
AU - Brudek, Tomasz
PY - 2016/11/17
Y1 - 2016/11/17
N2 - 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.
AB - 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.
U2 - 10.1038/srep37116
DO - 10.1038/srep37116
M3 - Journal article
C2 - 27853238
AN - SCOPUS:84995555368
VL - 6
JO - Scientific Reports
JF - Scientific Reports
SN - 2045-2322
M1 - 37116
ER -
ID: 178852308