Evaluation of Antifungal Activity of Defensin (Tfgd2) Using Its Heterologous Expression in E. coli

• Main Authors: Shiva Shiehbeiki, Mostafa Motallebi, Mohammad Zamani, Esmat Jourabchi
• Format: Article
• Language: English
• Published: University of Mazandaran 2021-02-01
• Series: Journal of Genetic Resources
• Subjects: antifungal activity; defensin; sytox green; tfgd2; pathogenesis-related protein
• Online Access: http://sc.journals.umz.ac.ir/article_3114_43897496dc534646c7199480c6253f97.pdf

Defensins are a superfamily of antimicrobial peptides that can inhibit the growth of a broad spectrum of fungi. To evaluate the antifungal activity of Tfgd2 gene of Trigonella foenum graecum, the coding region of this gene in the cDNA form was subcloned in the expression vector pET26b (+) and the construct was designated as pETSH1. The obtained construct expressing the recombinant protein with a hexahistidine tag at the C-terminal end transformed into the E. coli BL21 (DE3). Taguchi test applied for optimizing the protein expression and the expressed protein TFGD2 confirmed by SDS-PAGE and western blotting. The recombinant TFGD2 protein was purified using affinity chromatography with a Ni-NTA column. In vitro assay indicated a broad spectrum of antifungal activity of purified expressed recombinant TFGD2 against different fungal phytopathogens, such as Fusarium oxysporum, Rhizoctonia solani, Sclerotinia sclerotiorum, Alternaria solani, and Verticillium dahliae.