Isolation, Cloning, and in-silico Characterization of Glucanase (Glu) Gene/s From Trichoderma Spp.

Abstract

The present investigation entitled “Isolation, cloning and In-silico characterization of glucanase (Glu) gene/s from Trichoderma spp.” was carried out at department of Ag. Biotechnology SVPUAT, Meerut during 2019-2022. Glucanase gene produces hydrolytic enzymes such as β-1,3- glucanase, β-1,4-glucanase and β-1,6-glucanase that implicated in plant defense responses against fungal pathogens. These enzymes are responsible for breakdown of glucan, a major components of fungal cell walls. Genes encoding these enzymes have been used to genetically engineer plants to enhance their protection against fungal pathogens. The aim of this research was to isolates glucanolytic gene and to clone the genes encoding for glucanases with novel activities have been isolated from Trichoderma spp. of rhizospheric soil from different district of Uttar Pradesh. Seventeen species were assayed for antagonistic against Fusarium oxysporum and Sclerotium rolfsii, phytopathogenic fungi. The genomic DNA isolates from different species of Trichoderma, for glucanase gene amplification with specific primers. A novel β-1,6-glucanase gene from soil filamentous fungus Trichoderma harzianum (TBT-14) were isolated and cloned in pJET1.2/blunt cloning vector. The β-1,6- glucanase was 1328 bp long and encoded a 231 amino acid sequence that shared high identity with a β-1,6-glucanase from the TBT-14. The sequence contained conserved regions found in glycosyl hydrolases from family 5 that encode for substrate binding N-terminal sequences and putative asparagine linked glycosylation sites. The sequence of β-1,6-glucanase gene subjected to in-silico characterization and predict the protein structure.