PEG-Mediated Protoplast Transformation of Penicillium sclerotiorum (scaumcx01): Metabolomic Shifts and Root Colonization Dynamics
Year: 2025
Type: article
Abstract: Protoplast-based transformation is a vital tool for genetic studies in fungi, yet no protoplast method existed for P. sclerotiorum-scaumcx01 before this study. Here, we optimized protoplast isolation, regeneration, and transformation efficiency. The highest protoplast yield (6.72 × 106 cells/mL) was obtained from liquid mycelium after 12 h of enzymatic digestion at 28 °C using Lysing Enzymes, Yatalase, cellulase, and pectinase. Among osmotic stabilizers, 1 M MgSO4 yielded the most viable protoplasts. Regeneration occurred via direct mycelial outgrowth and new protoplast formation, with a 1.02% regeneration rate. PEG-mediated transformation with a hygromycin resistance gene and GFP tagging resulted in stable GFP expression in fungal spores and mycelium over five generations. LC/MS-based metabolomic analysis revealed significant changes in glycerophospholipid metabolism, indicating lipid-related dynamics influenced by GFP tagging. Microscopy confirmed successful colonization of tomato roots by GFP-tagged scaumcx01, with GFP fluorescence observed in cortical tissues. Enzymatic (cellulase) seed pretreatment enhanced fungal colonization by modifying root surface properties, promoting plant–fungal interaction. This study establishes an efficient protoplast transformation system, reveals the metabolic impacts of genetic modifications, and demonstrates the potential of enzymatic seed treatment for enhancing plant–fungal interactions.
Source: Journal of Fungi
DOI: 10.3390/jof11050386
Authors: Israt Jahan, Qilin Yang, Zijun Guan, Yihan Wang, Ping Li, Jian Yan
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
SDG: Zero hunger
Open Access status: gold
Estimated APC paid: $3,137
OpenAlex ID: W4410501675