Nicotiana benthamiana is used extensively as a platform for transient gene expression and a model system for studying plant-virus interactions. However, many tobacco-feeding insects, including Myzus persicae (green peach aphids) and Bemisia tabaci (whiteflies), grow poorly on N. benthamiana. Using CRISPR/Cas9, we generated knockout mutations in two N. benthamiana acylsugar acyltransferase genes, ASAT1 and ASAT2. Whereas ASAT1 mutations reduced the abundance of two predominant acylsucroses, ASAT2 mutations caused almost complete depletion of insect-deterrent acylsucroses. Both M. persicae and B. tabaci survived and reproduced better on asat2 mutant than on wildtype N. benthamiana plants. The improved aphid and whitefly performance on ASAT2 mutants make it easier to use the efficient transient overexpression and gene expression silencing systems that are available for N. benthamiana to study plant-insect interactions. Additionally, the absence of acylsugars in ASAT2 mutant lines will simplify transient expression assays for the functional analysis of acylsugar biosynthesis genes from other Solanaceae.
