Fig. 8
Analysis of the interaction between LrWRKY3 and LrDef1 promoter. a EMSA revealed that the LrWRKY3 specifically bound to W box. 1, reaction solution containing only free probes; 2, LrWRKY3 can interact with biotin labelled probes containing the W box sequence; 3, LrWRKY3 can both interact with biotin labelled probes containing the W box sequence and unlabeled probes containing the W box sequence; 4, LrWRKY3 can’t interact with mutant probes; Complex: the combination of LrWRKY3 recombinant protein and probes; Free probe: the unbound probes. b Analysis of the transactivation effect of LrWRKY3 on LrDef1 promoter. LrWRKY3 + pLrDef1: transcriptional activity analysis of pGADT7-LrWRKY3 interaction with pLrDef1; Positive control: pGADT7-p53 can interact with pAbAi-p53 as a positive control; Negative control: pAbAi empty vector was used a negative control. c GUS activity was higher in transgenic tobacco lines co-expressing of LrWRKY3 and LrDef1 promoter than in the transgenic tobacco lines expressing of LrDef1 promoter. The results were shown as average values calculated from three replicates and calculated by the 2-ΔΔCt method and analyzed by the Student’s t test. pBI121: pBI121-GUS transgenic tobacco; PD-2/3/7/10: pLrDef1 transgenic tobacco lines. Co-1/7/9/12: LrWRKY3/pLrDef1 transgenic tobacco lines