Advances in Zoology and Botany Vol. 5(2), pp. 17 - 24
DOI: 10.13189/azb.2017.050202
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Evaluation of Reference Genes for Differential Gene Expression Study of Bovine Tuberculosis

Ailam Lim 1,2,*, Steven Bolin 2
1 Breathitt Veterinary Center, Murray State University, United States
2 Department of Pathobiology and Diagnostic Investigation, Michigan State University, United States


Relative quantitative real-time PCR (qPCR) assays serve as important tools for validating differential gene expression data. A reference gene that is stably expressed across sample types and experimental treatments is crucial for accuracy in interpretation of relative qPCR data. Twelve previously validated reference genes were evaluated in this study to identify a most suitable reference gene that can be used for gene expression study of bovine tuberculosis (bTB) infected and bTB test-false positive cattle, in peripheral blood mononuclear cells after a 4 hour or after an overnight stimulation with bovine tuberculin antigen. Stability of the candidate reference genes were evaluated using the BestKeeper, geNorm and NormFinder programs. The SDHA was found to be the most stably expressed reference gene, regardless of infection status and varying length (4 hours or overnight) of antigen stimulation, while expression of many widely used reference genes are not stable under the studied experimental conditions. We also confirmed that the geNorm and NormFinder programs yielded similar findings in determining the stability of reference genes, which differ largely from the BestKeeper program. This finding stresses the importance of validating the reference gene(s) chosen for each experimental study, and the need for using multiple programs for the evaluation.

Reference Genes, Gene Expression, Quantitative Real-time PCR, Bovine Tuberculosis, Peripheral Blood Mononuclear Cells

Cite This Paper in IEEE or APA Citation Styles
(a). IEEE Format:
[1] Ailam Lim , Steven Bolin , "Evaluation of Reference Genes for Differential Gene Expression Study of Bovine Tuberculosis," Advances in Zoology and Botany, Vol. 5, No. 2, pp. 17 - 24, 2017. DOI: 10.13189/azb.2017.050202.

(b). APA Format:
Ailam Lim , Steven Bolin (2017). Evaluation of Reference Genes for Differential Gene Expression Study of Bovine Tuberculosis. Advances in Zoology and Botany, 5(2), 17 - 24. DOI: 10.13189/azb.2017.050202.