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A pathogen-inducible ethylene-responsive factor gene, designated as EREB5, was firstly isolated and characterized from G.barbadense (Gossypium barbadense L.) inoculated with V.dahliae (Verticillium dahliae).Bioinformatics analysis demonstrated that the deduced protein of 156 amino acid residues possessed a conserved ERF (Ethylene-response factor) domain, and some other typical features of transcription factors, including one nucleus localization signal, one serine-rich domain, several phosphorylation sites, and one motif of potato inhibitor I family.Transient expression analysis in onion epidermal cell showed that EREB5 protein was localized in nucleus.Meanwhile, gel shift assay revealed that EREB5 protein had the ability of binding specifically to the GCC-box.Moreover, the results of expression analysis indicated that EREB5 protein was able to be induced in G.barbadense seedlings, by inoculating with V.dahliae and stimulation of ethylene, respectively.In tobacco, overexpression of EREB5 activated pathogenesis-related genes under normal growth conditions.These results imply that EREB5 should play an important role in resistance of G.barbadense during the resistance response to Verticillum wilt.