Two Novel Defensin Genes from Brassica Juncea and Camelina Sativa Confers Antifungal Activity Against Pathogenic Fungi Alernaria brassicae
GDG Chaturani1*, Zahoor Ahmad Mir2 and Anita Grover3
1PhD Scholar, National Institute of Plant Biotechnology, New Delhi, India
2Senior Research Fellow, National Institute of Plant Biotechnology, New Delhi, India
3Principle Scientist, National Institute of Plant Biotechnology, New Delhi, India
*Corresponding Author: GDG Chaturani, PhD Scholar, National Institute of Plant Biotechnology, New Delhi, India.
November 02, 2021; Published: January 28, 2022
Alternaria blight, incited by A. brassicae is one of the most common and destructive fungal pathogens in Indian mustard which accounts for 46% of yield loss. Efforts are being made in order to have sufficient knowledge about the genes induced during infection and their regulation measures. We hear report two novel defensins, BjDef and CsDef which confers antifungal activity against A. brassicae. During the search of defensins, cloning and sequencing resulted in two open reading frames of 234 bp with characteristic α-helix, triple strand antiparallel β-sheets, highly conserved eight cysteines and an identical backbone structure stabilized by four intra-molecular disulphide bridges in their peptide chains. Amplified fragments were cloned into pET29a vector system and were expressed in Escherichia coli. Recombinant defensin proteins were optimally expressed in isopropyl thio-galactoside concentration of 1.0 mM/ml and 6 h of incubation. Purified proteins with hexa-histidine tag by Ni-NTA column exhibited a 16 kDa single fragment. The purified proteins showed an inhibitory effect on hyphal growth indicating disrupted mycelial cells while percentage mycelial growth inhibition calculations indicate inhibition effect of proteins in a concentration dependent manner. Even though at lower concentration inhibition was high by BjDef, at higher concentrations CsDef conferred comparatively more effective inhibition. Relative expression of CsDef revealed a high level of expression at 48 h in response to Alternaria infection. Comparatively low level of expression was reported by BjDef. Our results confirmed the ability of above defensins to be used in construction of transgenic brassica to confirm resistance to A. brassicae.
Keywords: Alternaria Blight; Brassica Juncea; Defensin; Antifungal
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