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Selection of Candidate Housekeeping Genes for Normalization in Human Postmortem Brain Samples
Oncology, Biology, and Genetics Department (DOBiG), University of Genoa, Largo R. Benzi, 10, 16132 Genoa, Italy
Department of Health Sciences, University of Molise, Via De Sanctis, 86100 Campobasso, Italy
National Institute for Cancer Research (IST) Genoa, Largo R. Benzi, 10, 16132 Genoa, Italy
These authors contributed equally to this work.
* Author to whom correspondence should be addressed.
Received: 17 June 2011; in revised form: 28 July 2011 / Accepted: 17 August 2011 / Published: 26 August 2011
Abstract: The most frequently used technique to study the expression profile of genes involved in common neurological disorders is quantitative real-time RT-PCR, which allows the indirect detection of very low amounts of selected mRNAs in tissue samples. Expression analysis by RT-qPCR requires an appropriate normalization to the expression level of genes characterized by a stable, constitutive transcription. However, the identification of a gene transcribed at a very stable level is difficult if not impossible, since significant fluctuations of the level of mRNA synthesis often accompanies changes of cell behavior. The aim of this study is to identify the most stable genes in postmortem human brain samples of patients affected by Alzheimer’s disease (AD) suitable as reference genes. The experiments analyzed 12 commonly used reference genes in brain samples from eight individuals with AD and seven controls. After a careful analysis of the results calculated by geNorm and NormFinder algorithms, we found that CYC1 and EIF4A2 are the best reference genes. We remark on the importance of the determination of the best reference genes for each sample to be analyzed and suggest a practical combination of reference genes to be used in the analysis of human postmortem samples.
Keywords: Alzheimer’s disease; quantitative real-time RT-PCR; reference genes; geNorm; NormFinder
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Penna, I.; Vella, S.; Gigoni, A.; Russo, C.; Cancedda, R.; Pagano, A. Selection of Candidate Housekeeping Genes for Normalization in Human Postmortem Brain Samples. Int. J. Mol. Sci. 2011, 12, 5461-5470.
Penna I, Vella S, Gigoni A, Russo C, Cancedda R, Pagano A. Selection of Candidate Housekeeping Genes for Normalization in Human Postmortem Brain Samples. International Journal of Molecular Sciences. 2011; 12(9):5461-5470.
Penna, Ilaria; Vella, Serena; Gigoni, Arianna; Russo, Claudio; Cancedda, Ranieri; Pagano, Aldo. 2011. "Selection of Candidate Housekeeping Genes for Normalization in Human Postmortem Brain Samples." Int. J. Mol. Sci. 12, no. 9: 5461-5470.