Selection of reference genes and expression of key genes involved in response to short-term soil flooding stress in Prunus persica rootstock ‘Tsukuba 1’

Abstract

The hypoxic environment directly impacts energy production, growth, and plant development. Under this condition, plants need to rely on anaerobic metabolism to maintain ATP production. These responses are modulated by complex molecular mechanisms that are responsible for gene expression to avoid plant damage. In order to carry out gene expression studies using the real-time PCR technique (RT-qPCR), it is necessary to normalize the data based on optimal reference gene(s) to experimental conditions. Thus, the first aim of this study was to identify likely reference genes and then determine their stability. Besides, we investigated whether adjusting root energy that ‘Tsukuba 1’ plays in the hypoxia is compatible with tolerance or susceptibility to short-term soil flooding. For that, ‘Tsukuba 1’ was submitted to two irrigation treatments: 1) non-flooded, and 2) flooded (1, 2, 4, 8, 16, and 32 h). A comprehensive analysis of performance and stability showed as most stable reference genes, PpTEF2 and PpTUA for roots and PpPLA2 and PpCYP2 for leaves. Conversely, PpTUB and PpEf1α, and PpTUA genes were not suitable reference genes for roots and leaves, respectively. Under short-term soil flooding, ‘Tsukuba 1’ roots generate energy anaerobically firstly through lactic-acid metabolism and then by alcoholic in 2 and 8 h, respectively. In summary, moderate tolerance to short-term soil flooding was observed in peach cultivar ‘Tsukuba 1’, mainly due to stimulation of glycolysis and up-regulation of key gene expression responsible for pivotal pathways involved in energy and ethylene metabolism. However, other physiological and metabolic traits/responses needed to be investigated to confirm this trend.