Selection of Reference Genes and Expression Profiling in Heat-Stressed Grapevine Varieties.

Background: "Touriga Franca" (TF) and "Touriga Nacional" (TN) are grapevine varieties cultivated in the 'Douro Superior' subregion (Northern Portugal) that experience stressful environmental conditions during the summer.

Objectives: Aiming to profile the expression of stress-responsive genes by quantitative real-time PCR (qPCR) in TF and TN plants growing naturally, three candidate reference genes were first tested under controlled conditions.

Methods: To simulate a summer's day, TF and TN in vitro plants were exposed to 32 °C-3 h (heat acclimation) and 42 °C-1 h (severe heat stress, HS) followed by two recovery periods (32 °C-3 h and 24 °C-24 h). Leaf samples were collected at the end of each phase. Control plants were kept at 24 °C.

Results: Among the candidate reference genes, the and pair showed the highest stability. The suitability of these genes for qPCR was validated by () gene profiling. The expression was up-regulated in both varieties and all experimental phases except in TF control plants. TN showed the highest relative expression ratio after severe HS.

Conclusions: TN responded faster than TF to the induced heat shocks. The , , and genes revealed to be suitable for further qPCR assays in TF and TN grapevine varieties.