Physiological drought resistance and accumulation of leaf phenolics in white clover interspecific hybrids

•First study of functional properties in novel hybrids for drought-exposed pastures.•The stress-resistant BC1 hybrids were able to maintain photosynthesis under drought.•Drought resistance in BC1 hybrids was related to high constitutive kaempferol levels.•BC1 hybrids also showed the highest increases of hydroxycinnamic acids under drought.•BC1 families with high levels of specific phenolics can be used for plant improvement. This study investigated drought responses in first (BC1) and second (BC2) backcross generation hybrids of Trifolium repens L.×T. uniflorum L., and compared these to high-yielding T. repens L. (white clover) cultivars. Plant attributes included accumulation of leaf phenolics as well as other biochemical and physiological water stress responses. Measurements were made across broad clover types as well as on a subset of related material. Under drought, net photosynthesis decreased by 44–48% and transpiration rates decreased by 60% in the BC2 family and in the white clover parent, but were unaltered in the BC1 family. Drought-induced decreases in leaf water potential were more pronounced in the BC1 family (−47%) than the BC2 (−31%) and parental (−28%) material. Quercetin glycoside accumulation generally increased 2–3 fold under drought. Compared with the BC2 family and with parent plants, the BC1 family had 2.5–4x higher constitutive kaempferol glycoside levels, and 1.5–2.5x higher kaempferol glycoside accumulation under drought. Constitutive kaempferol glycoside accumulation was related to reduced senescence and to less pronounced decreases in shoot DW under drought stress. Hydroxycinnamic acid accumulation increased by 56–73% under water stress, particularly for the BC1 generation. These results identify physiological and biochemical traits that help explain drought resistance of T. repens×T. uniflorum hybrids. The findings suggest merit for the selection of BC1 families with increased accumulation of protective phenolic compounds in breeding programmes for improved drought resistance.