Many Gram negative bacteria require a Type III secretion system (T3SS) to cause disease. Translocation of Type III-secreted effectors into host cells interferes with host immunity and modifies host physiology. For most pathosystems, researchers have focused on effector function, but we lack a spatial understanding of host cells targeted by Type III effectors. The pathogenic bacterium Xanthomonas translucens depends on a T3SS to cause bacterial leaf streak on barley. Type III secreted transcriptional activator-like (TAL) effectors found in X. translucens bind promoters of host genes and induce plant gene expression. We developed a tool to visualize cells specifically targeted by X. translucens TAL effectors. We engineered transgenic barley that only express green fluorescent protein (GFP) in the presence of TAL effectors translocated by X. translucens. We created artificial, designer TAL effectors (dTALEs) that specifically bind the the promoter of the GFP gene. We confirmed through quantitative PCR that specifically GFP was only induced by X. translucens in a TAL effector-dependent manner. With fluorescence laser confocal microscopy we found that the mesophyll and stomatal cells were targeted by X. translucens during leaf infection. We expect this tool to bring new insights in TAL effector-targeting of host cells during barley infection by X. translucens.