Rhizobium rhizogenes A4-derived strains mediate hyper-efficient transient gene expression in Nicotiana benthamiana and other solanaceous plants

Agroinfiltration, a method using agrobacteria to transfer DNA into plant cells, is widely used for transient gene expression in plants.Besides the commonly used Agrobacterium strains, Rhizobium rhizogenes can also deliver foreign DNA into host plants.While many R. rhizogenes strains have been known for inducing hairy root symptoms, their use for transient expression has not been fully explored.Here, we showed that R. rhizogenes A4 outperformed all other tested agrobacterial strains in agroinfiltration assays on leaves of Nicotiana benthamiana and other solanaceous plants.By conducting an agroinfiltration screening in N. benthamiana using various agrobacterial strains carrying the RUBY reporter, we discovered that A4 mediates the strongest and fastest transient expression.Utilizing the genomic information, we developed a collection of disarmed and modified strains derived from A4.By performing vacuum infiltration assays, we showed that these A4-derived strains efficiently transiently transform 6-week-old N. benthamiana leaves compared to the GV3101 strain.Furthermore, we performed agroinfiltration using the A4-derived strain AS109 on the leaves of tomato, pepper, and eggplant.Remarkably, AS109 mediated transient gene expression more effectively than all tested commonly used strains.This discovery paves the way for establishing A4-derived strains for enhancing transient expression in N. benthamiana and facilitating functional studies in other plant species.