Reference Gene Selection and Prednisolone Target Gene Expression in Adipose Tissues of Friesian Cattle

Aspergillus is a genus of filamentous fungi with vast geographic and ecological distributions. Species within this genus are clinically, agriculturally and biotechnologically relevant, leading to increasing interest in elucidating gene expression dynamics of key metabolic and physiological processes. Reverse-transcription quantitative Polymerase Chain Reaction (RT-qPCR) is a sensitive and specific method of quantifying gene expression. A crucial step for comparing RT-qPCR results between strains and experimental conditions is normalisation to experimentally validated reference gene(s). In this review, we provide a critical analysis of current reference gene selection and validation practices for RT-qPCR gene expression analyses of Aspergillus. Of 90 primary research articles obtained through our PubMed query, 17 experimentally validated the reference gene(s) used. Twenty reference genes were used across the 90 studies, with beta-tubulin being the most used reference gene, followed by actin, 18S rRNA and glyceraldehyde 3-phosphate dehydrogenase. Sixteen of the 90 studies used multiple reference genes for normalisation. Failing to experimentally validate the stability of reference genes can lead to conflicting results, as was the case for four studies. Overall, our review highlights the need to experimentally validate reference genes in RT-qPCR studies of Aspergillus.

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Optimisation of region-specific reference gene selection and relative gene expression analysis methods for pre-clinical trials of Huntington's disease

Molecular Neurodegeneration ◽

10.1186/1750-1326-3-17 ◽

2008 ◽

Vol 3 (1) ◽

pp. 17 ◽

Cited By ~ 36

Author(s):

Caroline L Benn ◽

Helen Fox ◽

Gillian P Bates

Keyword(s):

Gene Expression ◽

Clinical Trials ◽

Reference Gene ◽

Expression Analysis ◽

Gene Expression Analysis ◽

Gene Selection ◽

Specific Reference ◽

Relative Gene Expression ◽

Reference Gene Selection ◽

Relative Gene

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Reference gene selection for qPCR gene expression analysis of rust-infected wheat

Physiological and Molecular Plant Pathology ◽

10.1016/j.pmpp.2012.10.006 ◽

2013 ◽

Vol 81 ◽

pp. 22-25 ◽

Cited By ~ 34

Author(s):

Jakobus J. Scholtz ◽

Botma Visser

Keyword(s):

Gene Expression ◽

Reference Gene ◽

Expression Analysis ◽

Gene Expression Analysis ◽

Gene Selection ◽

Reference Gene Selection ◽

Selection For

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Validation of internal reference genes for relative quantitation studies of gene expression in human laryngeal cancer

PeerJ ◽

10.7717/peerj.2763 ◽

2016 ◽

Vol 4 ◽

pp. e2763 ◽

Cited By ~ 3

Author(s):

Xiaofeng Wang ◽

Jinting He ◽

Wei Wang ◽

Ming Ren ◽

Sujie Gao ◽

...

Keyword(s):

Gene Expression ◽

Reference Gene ◽

Laryngeal Cancer ◽

Reference Genes ◽

Target Gene ◽

Cancer Tissue ◽

Internal Reference ◽

Target Gene Expression ◽

Internal Reference Gene ◽

Relative Quantitation

BackgroundThe aim of this study was to determine the expression stabilities of 12 common internal reference genes for the relative quantitation analysis of target gene expression performed by reverse transcription real-time quantitative polymerase chain reaction (RT-qPCR) in human laryngeal cancer.MethodsHep-2 cells and 14 laryngeal cancer tissue samples were investigated. The expression characteristics of 12 internal reference gene candidates (18S rRNA, GAPDH, ACTB, HPRT1, RPL29, HMBS, PPIA, ALAS1, TBP, PUM1, GUSB, and B2M) were assessed by RT-qPCR. The data were analyzed by three commonly used software programs: geNorm, NormFinder, and BestKeeper.ResultsThe use of the combination of four internal reference genes was more appropriate than the use of a single internal reference gene. The optimal combination was PPIA + GUSB + RPL29 + HPRT1 for both the cell line and tissues; while the most appropriate combination was GUSB + RPL29 + HPRT1 + HMBS for the tissues.ConclusionsOur recommended internal reference genes may improve the accuracy of relative quantitation analysis of target gene expression performed by the RT-qPCR method in further gene expression research on laryngeal tumors.

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