Optimization of transient Agrobacterium-mediated gene expression system in leaves of Nicotiana benthamiana

Here we report on a simple and reproducible system of Agrobacterium-mediated transient gene expression assay that utilizes infiltration of young Nicotiana benthamiana leaves. Although some of the phenomena described in this paper have been already reported by other researchers, here we have further developed them. The highest level of transient gfp gene expression was detected in the youngest leaves of N. benthamiana infiltrated with A. tumefaciens strains AGL0 and EHA105 precultured in the presence of 450-600 µM acetosyringone. Although the maximum level of transient gfp gene expression was restricted presumably by RNA silencing, it was completely suppressed in the presence of the viral protein HC-Pro. The transient expression system described here can be used to identify new viral suppressors of RNA silencing, for detailed analysis of unidentified genes and for industrial production of proteins in plants as well.