Application progress of CRISPR/Cas9 genome-editing technology in edible fungi

Abstract

Edible fungi are not only delicious but are also rich in nutritional and medicinal value, which is highly sought after by consumers. As the edible fungi industry continues to rapidly advance worldwide, particularly in China, the cultivation of superior and innovative edible fungi strains has become increasingly pivotal. Nevertheless, conventional breeding techniques for edible fungi can be arduous and time-consuming. CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats/CRISPR-associated nuclease 9) is a powerful tool for molecular breeding due to its ability to mediate high-efficiency and high-precision genome modification, which has been successfully applied to many kinds of edible fungi. In this review, we briefly summarized the working mechanism of the CRISPR/Cas9 system and highlighted the application progress of CRISPR/Cas9-mediated genome-editing technology in edible fungi, including Agaricus bisporus, Ganoderma lucidum, Flammulina filiformis, Ustilago maydis, Pleurotus eryngii, Pleurotus ostreatus, Coprinopsis cinerea, Schizophyllum commune, Cordyceps militaris, and Shiraia bambusicola. Additionally, we discussed the limitations and challenges encountered using CRISPR/Cas9 technology in edible fungi and provided potential solutions. Finally, the applications of CRISPR/Cas9 system for molecular breeding of edible fungi in the future are explored.

Current Advances in the Functional Genes of Edible and Medicinal Fungi: Research Techniques, Functional Analysis, and Prospects

“…However, the CRISPR/Cas9 system faces several challenges, including low editing efficiency, poor accuracy, off-target effects, and difficulties in screening mutants [95]. In the future, we can improve the editing efficiency by optimizing the codons of target species and selecting appropriate promoters to express Cas9 genes; improve the accuracy by inhibiting NHEJ repair or enhancing HDR repair; detect off-target effects on time using chromatin immunoprecipitation sequencing (ChIP-seq) and genome-wide, unbiased identification of DSBs enabled by sequencing (GUIDE-seq) methods, optimized sgRNA sequences, and the modification of Cas9 proteins to reduce off-target effects; and the addition of chemical reagents to increase the proportion of mononuclear protoplasts to improve the efficiency of obtaining pure transformants, thus reducing the difficulty of screening [96].…”

Advances in biosynthesis and metabolic engineering strategies of cordycepin

“…Cordyceps genus undergo a complex two-stage growth process and face high rates of fruit-body gemmated degeneration. Above factors contribute to additional difficulties in implementing the CRISPR system in these species ( Zhang et al, 2023 ). In C. militaris , the pyrG gene was successfully edited using CRISPR/Cas9 system, with the efficiency of 11.76%.…”