Evaluation of candidate reference genes for RT-qPCR in lily (Lilium brownii)

Abstract

SummaryThe selection of a reliable reference gene (or genes) is essential for correct interpretation of real-time reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) data. Thus, eight frequently-used reference genes in lily (Lilium brownii), were selected to evaluate their stability of expression under various treatments including the application of plant growth regulators (PGRs) or stress (including cold, heat, drought, or heavy metals) and in various tissues including roots, stems, leaves, sepals, petals, stamens, and pistils. Based on a comprehensive analysis using geNorm and NormFinder software, elongation factor 1-alpha (EF1α) and the gene for 18S rRNA (18S rRNA) emerged as the most stable reference genes when the data from all 23 lily samples were combined. The genes for glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and photosystem I P700 apoprotein A (psaA) could serve as reliable internal reference genes in different tissues, while psaA and 18S rRNA ranked top in stressed leaves and in PGR-treated roots. GAPDH and EF1α exhibited the most stable expression patterns in leaves treated with PGRs, while psaA and EF1α were the optimal internal reference genes in stressed roots. On the other hand, the RNA polymerase C gene (RpoC1) and the ATP synthase β-subunit gene (ATPB) were the least stable reference genes, and should be used with caution as reference genes during RT-qPCR analysis in lily.