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Fig. 6 | BMC Plant Biology

Fig. 6

From: Inhibition of multiple defense responsive pathways by CaWRKY70 transcription factor promotes susceptibility in chickpea under Fusarium oxysporum stress condition

Fig. 6

CaWRKY70 suppresses CaWRKY40 promoter activity in vivo and in planta. a In vivo Chromatin immunoprecipitation (ChIP) PCR assay shows CaWRKY70 binding to W-box 2 of CaWRKY40 promoter at shoot tissues of susceptible (JG62) chickpea under Foc1 stress. Diagram shows presence of W-boxes at CaWRKY40 promoter. (In) denotes input amplified from pre-cleared chromatin samples. Arrow indicates position of the primers. Plus (+) and minus (−) signs indicate anti-CaWRKY70 antibody and pre-immune sera immunoprecipitated chromatins. (+ 1) denotes transcription start site (TSS). b In silico molecular docking of CaWRKY70 and W-box 2 containing CaWRKY40 promoter DNA of cluster 3. c, d Electrophoretic mobility shift assay (EMSA) shows in vitro histidine tagged WRKY70 binding at pWRKY40 W-box 2. Approximately, 200 ng of WRKY70-His protein specifically binds at W-box 2 (− 217 to − 245 bp upstream of TSS). Plus (+) and minus (−) signs indicate presence or absence of specific components. BP indicates bound probe and FP represents free probe. Box indicates W-box 2. Asterisk (*) indicates mutated W-box. The experiment was repeated twice with similar results. e CaWRKY70 mediated trans-inhibition of CaWRKY40 promoter activity. p35S:CaWRKY70 and pWRKY40:YFP constructs were co-transfected in protoplasts obtained from Nicotiana tabacum cv. Xanthi. (Brad) cell suspension culture. mCherry was used as transformation marker. Scale bar = 10 μm. (f) CaWRKY70 reduces GUS expression driven by CaWRKY40 promoter in tobacco leaf discs. Diagram shows constructs used for Agrobacterium mediated transient co-infiltration experiment. Plus (+) and minus (−) signs indicate presence or absence of specific vehicles

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