Abstract Pleurotus ostreatus is one of the most commercially produced edible mushrooms worldwide. Improved cultivated strains with more useful traits have been obtained using classical breeding, which is laborious and time-consuming. Here, we attempted efficient gene mutagenesis using plasmid-based CRISPR/Cas9 as the first step for non-genetically modified (non-GM) P. ostreatus generation. Plasmids harboring expression cassettes of Cas9 and different single guide RNAs targeting fcy1 and pyrG were individually transferred into fungal protoplasts of the PC9 strain, which generated some strains exhibiting resistance to 5-fluorocytosine and 5-fluoroorotic acid, respectively. Genomic PCR followed by sequencing revealed small insertions/deletions or insertion of a fragment from the plasmid at the target site in some of the drug-resistant strains. The results demonstrated efficient CRISPR/Cas9-assisted genome editing in P. ostreatus , which could contribute to the molecular breeding of non-GM cultivated strains in the future. Furthermore, a mutation in fcy1 via homology-directed repair using this CRISPR/Cas9 system was also efficiently introduced, which could be applied not only for precise gene disruption, but also for insertions leading to heterologous gene expression in this fungus.

Tatpong Boontawon

Takehito Nakazawa

Chikako Inoue

Keishi Osakabe

Moriyuki Kawauchi

Masahiro Sakamoto

Yoichi Honda

AMB Express

AMB EXPRESS 是一本高质量的期刊，汇集了应用和工业微生物学领域的研究，尤其对“白色生物技术”和“红色生物技术”感兴趣。重点是使用微生物、真核细胞培养物或酶进行化合物生物合成、转化和降解的过程。这包括精细和散装化学品、聚合物和酶或其他蛋白质。还考虑了下游过程。还公布了结合生化和化学过程的综合过程。

AMB Express is a high-quality journal that focuses on research in the fields of applied and industrial microbiology, with particular interest in 'white biotechnology' and 'red biotechnology.' The journal emphasizes the use of microorganisms, eukaryotic cell cultures, or enzymes for the biosynthesis, transformation, and degradation of compounds. This includes studies on fine and bulk chemicals, polymers, and enzymes or other proteins. The journal also considers downstream processes and publishes integrated research that combines biochemical and chemical processes.

AMB Express 是一本高质量的期刊，专注于应用和工业微生物学领域的研究，特别关注“白色生物技术”和“红色生物技术”。期刊的重点是利用微生物、真核细胞培养物或酶进行化合物的生物合成、转化和降解。这包括精细化学品、散装化学品、聚合物以及酶或其他蛋白质的相关研究。同时，期刊也关注下游过程，并发布结合生化和化学过程的综合研究成果。

Efficient genome editing with CRISPR/Cas9 in Pleurotus ostreatus

Efficient CRISPR/Cas9-assisted genome editing is demonstrated in P. ostreatus, which could contribute to the molecular breeding of non-GM cultivated strains in the future and could be applied not only for precise gene disruption, but also for insertions leading to heterologous gene expression in this fungus.

Category	Quartile
生物学	3
生物学, 生物工程与应用微生物	3

Efficient genome editing with CRISPR/Cas9 in Pleurotus ostreatus

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Journal Info

Category	Quartile
BIOTECHNOLOGY & APPLIED MICROBIOLOGY	2