Wang et al. 16135 Figure 9. Tobacco plants challenged with Black Shank P. parasitica. Pictures were taken eight days after P. palmivora inoculation. a, c, e, g and i show transgenic plants, while b, d, f, h and j, show controls, the T1 plants transformed with plasmid pBI121. Compared with control, the transgenic tobacco plants were still green after pathogen infection. over bacterial expression systems, is that the yeast has the potential to perform many of the post-translational modifications typically associated with higher eukaryotes, such as the processing of signal sequences, folding, disulfide bridge formation, certain types of lipid addition and glycosylation (Cereghino and Cregg, 2000). Eight cysteine residues present in ZmDEF1 mature protein play an important role in protein stabilization by formation multiple disulfide bridges. P. pastoris is believed to be more effective in promoting disulfide bonding than the
16136 Afr. J. Biotechnol. Escherichia coli (Cregg et al., 1993). In our previous work, we found that the fusion ZmDEF1 expressed <strong>using</strong> a prokaryotic system showed no antifungal activity (data not shown) possibility due to improper folding of the recombinant protein. However, the recombinant ZmDEF1 peptide created <strong>using</strong> the P. pastoris expression system exhibits an inhibitory activity on P. parasitica growth (Figure 6). Due to an alarming increase of resistance of microorganisms to classical antibiotics, the introduction and expression of antimicrobial peptides like plant defensins in crops is emerging as an intriguing biotechnological application for enhancing disease resistance (Punja, 2001; Osusky et al., 2000; Jha and Chattoo, 2010). In this work, a new plant defensin gene, ZmDEF1, was introduced into tobacco by Agrobacteriummediated transformation. Results indicate that constitutive expression of the ZmDEF1 gene under the control of the CaMV 35S promoter in tobacco results in enhanced resistance against P. parasitica (Figures 8 and 9). Thus, the antifungal activity of the defensin ZmDEF1 and its availability for genetic engineering should make it useful as gene source for engineering transgenic plants resistant against phytopathogenic fungi. ACKNOWLEDEGMENTS This work was supported by National Natural Science Foundation (Grant No. 31171258) and the projects (No. 2008ZX08009-003 and 2008ZX08003-002) <strong>from</strong> the Ministry of Agriculture of China for Transgenic Research. REFERENCES Almeida MS, Cabral KM, Zingali RB, Kurtenbach E (2000). Characterization of Two Novel Defense Peptides <strong>from</strong> Pea (Pisum sativum) Seeds. Arch. Biochem. Biophys. 378: 278-286. Aluru M, Curry J, O’Connell MA (1999). Nucleotide sequence of a defensin or-thionin-like gene (accession no. 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