Reference gene selection for quantitative real-time polymerase chain reaction in Populus.

Accurate quantification of gene expression with quantitative real-time polymerase chain reaction (qRT-PCR) relies on the choice of an appropriate reference gene. In this study, nine candidate reference genes were selected to study the expression stability for qRT-PCR normalization in adventitious rooting of Populus hardwood cuttings. geNorm, NormFinder, and BestKeeper analysis revealed that actin isoform B (ACT) was the most unstable gene across developmental stages, whereas elongation factor 1 alpha (EF1a) and 18S recombinant RNA (18S) emerged as the most appropriate reference genes for qRT-PCR analysis in this complex developmental process.