Journal
JOURNAL OF IMMUNOLOGICAL METHODS
Volume 384, Issue 1-2, Pages 196-199Publisher
ELSEVIER SCIENCE BV
DOI: 10.1016/j.jim.2012.07.020
Keywords
Lymph nodes; Lymphocytes; Reference genes; Quantitative PCR (qPCR)
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Funding
- National Institute of Health [NCI R01 CA68328, T32 CA080416]
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Normalization to a reference gene is the method of choice for quantitative PCR analysis. The stability of reference genes is critical for accurate gene expression analysis, as significant variations in reference gene expression can alter experimental results and conclusions. In this study, we evaluated the expression stability of five commonly used reference genes found in mouse lymphocytes. Using NormFinder and BestKeeper algorithms, we consistently show that ubiquitin C (Ubc) is the optimal reference gene for normalizing qPCR data obtained from mouse lymphocytes, whereas beta-actin (Actb) is not a suitable reference gene due to its extensive variability in expression. Our findings emphasize the importance of validating reference genes for qPCR analyses. We provide a shortlist of reference genes to use for normalization and recommend freely available software programs as a rapid approach to validate potential reference genes. (c) 2012 Elsevier B.V. All rights reserved.
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