Illumina NovaSeq 6000 sequencing; Rhizogenic Agrobacterium protein RolB interacts with the TOPLESS and NINJA repressor proteins to reprogram plant jasmonate signaling.
Rhizogenic Agrobacterium protein RolB interacts with the TOPLESS and NINJA repressor proteins to reprogram plant jasmonate signaling.
Collection of tomato roots infected with the wild-type K599 strain, the rolB∆ mutant strain and the rolB∆ strain with the constructs expressing either the native rolB oncogene or the rolBmEAR2 mutant. Tomato composite plant bioassays were carried out as previously described with some adaptations (Ho-Plágaro et al, 2018). More specifically, the WT rhizogenic Agrobacterium K599 (NCPPB2659; pRi2659) strain and the K599 strain mutated in the rolB oncogene through base editing, were used to infect tomato seedlings (Rodrigues et al, 2021). Both K599 strains were transformed as previously described with a plasmid expressing the mCherry fluorescent reporter under the control of the constitutive CaMV35S promoter from its T-DNA (Wen-Jun & Forde, 1989). In several experiments, the mCherry reporter construct was replaced by the pK7WG2D Gateway-compatible binary destination vector expressing a gene of interest under the control of the constitutive CaMV35S promoter and the GFP fluorescent marker under the control of the pRolD promoter from its T-DNA region. Per strain, ~30 tomato seedlings were infected as previously described (Ho-Plágaro et al, 2018). Four weeks post-infection, the roots were used for experimental purposes. Tomato plants were grown in a growth chamber under 16h/8h light/darkness photoperiod at 24°C for four weeks. Per strain, ~30 tomato seedlings were infected as previously described (Ho-Plágaro et al, 2018). RNA was extracted from root tissue with the RNeasy Plant Mini Kit (Qiagen). Library preparation was done using the TruSeq stranded RNA Sample Preparation Kit v2 (Illumina).