Fig. 5
Transient gene expression assays on N. benthamiana and barley plants. a Cell death was induced by candidate clones on the upper and lower surface of leaves on N. benthamiana plants as indicated. The Agrobacterium suspensions in 10 mM MgCl2 (OD600 > 0.5) were infiltrated with a needleless syringe into leaves of N. benthamiana plants, and the infiltrated plants were incubated at 24 °C in a growth room with a 16-h light/8-h dark cycle. Photos were taken at 5 days after infiltration. b Chlorosis was induced on barley leaves by transient expression of candidate genes without (Mock) and with (P131) the inoculation of the blast fungus strain P131. The β-glucuronidase (GUS) and LOC4331546 represent proteins incapable of inducing chlorosis. LOC4330898 and LOC9267830 represent proteins capable of triggering leaf chlorosis without or with subsequent blast fungus inoculation, respectively. Detached barley leaves transiently expressing candidate genes were inoculated with blast fungus strain P131 (5 × 105 spores/mL) and incubated in a growth chamber at 26 °C with a photoperiod of 16-h light/8-h darkness. Representative leaves were photographed at 96 h after inoculation. The experiment was repeated three times and similar results were observed. “+++” indicates tissue collapse of the Agrobacteria-inoculated region; “++” and “+” indicate partial or sporadic cell death, respectively, observed on the lower surface of leaves