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Fungal diseases are among the most important biotic stresses causing considerable losses in agricultural products. Improving crop plants with broad resistance to fungal diseases is one of the main challenges in agriculture. Classical plant breeding is too timely and has many other limitations, hence using biotechnology for this purpose is inevitable. Expression of a complex of Pathogen Related proteins induces polygenic resistance in plants. This technique leads to a broad and stable resistance to fungal pathogens. Hence, in this study we isolated resistance genes and constructed treble plasmid vectors with multiple genes involving three important families of disease-related proteins namely PR1, PR2, and PR3. For this purpose we isolated two genes of PR1 family with special primer design from tobacco genome, first. After analysis the accuracy of gene isolation using 35S promoter and Nos terminator, they were cloned in binary vector pBI121. Then, we cloned two important antifungal genes including PR2 (glucanase) and PR3 (chitinase) under the control of regulatory independent regions in T-DNA of the above mentioned vector with parallel and antiparallel orientations. These treble vectors have good antibacterial potential too and can be used for resistance to bacteria and other biotic and abiotic stresses. They can be introduced into plants using particle gun or through Agrobacterium mediated transformation.