Identification and validation of new reference genes for accurate quantitative reverse transcriptase-PCR normalization in the Antarctic plant Colobanthus quitensis under abiotic stress conditions

Abstract

The Antarctic ecotype of Colobanthus quitensis is a vascular plant highly adapted to the harsh environmental conditions of Maritime Antarctica which is now facing with the rapid local warming experienced in the Antarctic Peninsula during the last decades. Thus, the identification of the molecular mechanisms leading to the adaptation to this warming trend is a new target for modern cell physiology. The selection of suitable reference genes for quantification of key stress-responsive genes through quantitative Reverse Transcriptase-Polymerase Chain Reaction (qRT-PCR) is important to ensure accurate and reliable results. In this study, we evaluated the expression stability of eleven candidate genes in C. quitensis under different abiotic stress conditions using geNorm and RefFinder tools. The statistical analysis showed that the appropriate reference genes varied depending on the experimental conditions, even if EF1α and PP2Acs ranked as the most stable reference genes when all stress conditions were considered. To further validate the stability of the selected reference genes, the expression patterns of C. quitensis catalase gene (CqCAT) was analyzed. The reference genes validated in this study will be useful for improving the accuracy of qRT-PCR analysis for gene expression studies of the Antarctic ecotype of C. quitensis and could be extended to other ecotypes adapted to low temperatures.