Stability of housekeeping gene expression in Arabidopsis thaliana seedlings under differing macronutrient and hormonal conditions

Recent progress in plant hormone and macronutrient research is largely associated with characterising the molecular mechanisms of their actions. Quantitative reverse transcription-PCR is a powerful technique that enables elucidation of regulatory relationships through quantifying transcript levels, but its reliability requires precise normalization. We compared eight Arabidopsis thaliana housekeeping genes for expression stability after treatment with cytokinins, auxins, ethylene and after macronutrient level manipulation. The expression stability of housekeeping genes in cytokinin series was studied in wide spectrum of plants treated with different adenine- and phenylurea-type cytokinins as well as in plants with controlled overexpression of agrobacterial isopentenyltransferase. NormFinder and geNorm software were used to predict a unique set of genes as best for particular hormone groups. Generally, 18S rRNA, GAPC, and ACT were confirmed as less stable; and UBQ10, UBC, and EF-1α exhibited increased stability. Based on geNorm pairwise variation analysis, we confirmed accurate normalization required at least three reference genes.