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Title: Evaluation of Quantitative PCR Reference Genes for Gene Expression Studies in Tribolium castaneum After Fungal Challenge

Author
item Lord, Jeffrey
item Hartzer, Kris
item Toutges, Michelle
item Oppert, Brenda

Submitted to: Journal of Microbiological Methods
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/12/2009
Publication Date: 1/25/2010
Citation: Lord, J.C., Hartzer, K.L., Toutges, M.J., Oppert, B.S. 2010. Evaluation of Quantitative PCR Reference Genes for Gene Expression Studies in Tribolium castaneum After Fungal Challenge. Journal of Microbiological Methods. 80: 219-221.

Interpretive Summary: Quantitative polymerase chain reaction (qPCR) is the predominant means for measuring gene expression in all organisms and is widely used with the red flour beetle, a model insect for diverse biological investigations. Selection and validation of more than one stable reference gene is a basic but neglected requirement for qPCR. It is especially difficult when insects are heavily infected with pathogens that cause the breakdown of normal life processes. We have tested several candidate genes and identified three that are stable through several stages of larval development and at early and late stages of fungal infection and three that have acceptable stability. These reference genes will allow accurate quantification of gene expression in the red flour beetle and have broad applicability for beetles under stressful experimental conditions.

Technical Abstract: To investigate gene expression in Tribolium castaneum exposed to Beauveria bassiana, reference genes for qPCR were evaluated. Of these, the widely used genes for ß-actin, a-tubulin, and RPS6 were not stable. The most stable were ribosomal protein genes, RPS3, RPS18, and RPL13a. Syntaxin1, syntaxin6, and E-cadherin may be appropriate for some experimental systems.