Fig. 2: Engineered A. tumefaciens strains expressing a T3SS and AvrPto increase the transient and stable transformation efficiency of Arabidopsis.
a, b Transient transformation assay. a GUS staining of Arabidopsis leaves infiltrated with A. tumefaciens strain EHA105 (A600 = 0.2) harboring the binary vector pCAMBIA1301 (harbors GUS gene within the T-DNA) alone or in combination with pLN18, an empty vector, or a plasmid expressing AvrPto. Four days after infiltration, the leaves were stained with X-gluc and photographed. b Quantitative fluorometric GUS assays with extracts from leaves of plants treated as in a. c, d Root tumor assay. c Arabidopsis root segments were infected with the tumorigenic A. tumefaciens strain A208 (A600 = 0.01) expressing AvrPto in combination with or without pLN18. A. tumefaciens strains A208 and A208 (vector, pLN18) were also included as negative controls. Photographs were taken 4 weeks after A. tumefaciens infection. d Root segments forming tumors were counted from the experiment in c and the percentage of root segments forming tumors was calculated. e Root callus assay. Arabidopsis root segments were infected with non-tumorigenic A. tumefaciens strain EHA105 harboring the binary vector pCAS1 (A600 = 0.001) expressing AvrPto in combination with or without pLN18. A. tumefaciens strains EHA105 (pCAS1) and EHA105 (pCAS1, empty vector, pLN18) were included as negative controls. Four weeks after infection, root segments forming phosphinothricin (PPT)-resistant calli were counted and the percentage of root segments forming PPT-resistant calli was calculated. Data presented in b and d are mean ± standard error of three replicates. Bars with different letters are significantly different based on Tukey’s post-hoc one-way ANOVA analysis (p < 0.05). Brown-Forsythe test was done to test for variance (p = 0.6830 for b, 0.1433 for d). Data presented in e are mean of two replicates. Bars with different letters are significantly different based on Tukey’s post-hoc two-way ANOVA analysis (p = 0.0102). Experiments were repeated three times with similar results. Source data are provided as a Source Data file.
