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In nature, fungi play an important role in nutrient cycling, such as decomposition of dead organic matter ( Molina et al., 1993 Keizer, 1998 Pilz and Molina, 2001) and act as biofertilizers ( Hunt, 1999 Gates et al., 2005). We also discuss the potential and advantages of using targeted genome engineering in lichenized fungal (mycobiont) cultures to enhance their growth and secondary metabolite production in vitro can be complemented by other molecular approaches.įungal products are important in agriculture, textile, food, pharmaceutical industries, ( Cowan, 2001 Chang and Miles, 2004) and in bioremediation. Here, we discuss the recent research breakthroughs and developments which utilize CRISPR/Cas9 in the metabolic engineering of free-living fungi for the biosynthesis of secondary metabolites, enzyme production, antibiotics and to develop resistance against post-harvest browning of edible mushrooms and fungal pathogenesis. Artificially engineered molecular scissors, zinc finger nucleases (ZFNs), Transcriptional activator-like effector nucleases (TALENs that use protein motifs for DNA sequence recognition in the genome) and CRISPR associated protein 9 (Cas9 CRISPR/Cas9) system (RNA-DNA recognition) are being used in achieving targeted genome modifications for modifying traits in free-living fungal systems. These tools and the data generated are crucial for precise generation of fungal products such as enzymes, secondary metabolites, antibiotics etc. Studies using whole genome sequencing, computational and gene expression, targeted genome engineering techniques for generating site-specific sequence alterations through non-homologous end joining (NHEJ) by genomic double-strand break (DSB) repair pathway with high precision, resulting in gene inactivation have elucidated the complexity of gene expression, and metabolic pathways in fungi.
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