Transgenic Tobacco Plants Constitutively Overexpressing a Rice Thaumatin-like Protein (PR-5) Show Enhanced Resistance to Alternaria alternata

Abstract

Overexpression of antifungal pathogenesis-related (PR) proteins in crop plants has the potential for enhancing resistance against fungal pathogens. Thaumatin-like proteins (TLPs) are one group (PR-5, permatins) of antifungal PR-proteins isolated from various plants. In the present study, a plasmid containing a cDNA of rice tlp (D34) under the control of the CaMV-35S promoter was introduced into tobacco plants through Agrobacterium-mediated transformation system. A considerable overproduction of TLP was observed in transformed tobacco plants by Western blot analysis. There was a large accumulation of tlp mRNA in transgenic plants as revealed by Northern blot analysis. Southern blot analysis of the DNA from transgenic tobacco plants confirmed the presence of the rice tlp gene in the genomic DNA of transgenic tobacco plants. Immunoblot analysis of intracellular and extracellular proteins of transgenic tobacco leaves using a Pinto bean TLP antibody demonstrated that the 23-kDa TLP was secreted into the extracellular matrix. T2 progeny of regenerated plants transformed with TLP gene were tested for their disease reaction to Alternaria alternata, the brown spot pathogen. Transgenic tobacco plants expressing TLP at high levels showed enhanced tolerance to necrotization caused by the pathogen.