Development of a versatile and conventional technique for gene disruption in filamentous fungi based on CRISPR-Cas9 technology

Abstract Filamentous fungi represent an invaluable source of pharmaceutically active compounds. The development of versatile methods to genetically manipulate filamentous fungi is of great value for improving the low yields of bioactive metabolites and expanding chemical diversity. The CRISPR-Cas9-b...

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Autores principales:	Yan-Mei Zheng, Fu-Long Lin, Hao Gao, Gen Zou, Jiang-Wei Zhang, Gao-Qian Wang, Guo-Dong Chen, Zhi-Hua Zhou, Xin-Sheng Yao, Dan Hu
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2017
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/435d45484750440791e74b5b38caee94
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spelling	oai:doaj.org-article:435d45484750440791e74b5b38caee942021-12-02T11:41:20ZDevelopment of a versatile and conventional technique for gene disruption in filamentous fungi based on CRISPR-Cas9 technology10.1038/s41598-017-10052-32045-2322https://doaj.org/article/435d45484750440791e74b5b38caee942017-08-01T00:00:00Zhttps://doi.org/10.1038/s41598-017-10052-3https://doaj.org/toc/2045-2322Abstract Filamentous fungi represent an invaluable source of pharmaceutically active compounds. The development of versatile methods to genetically manipulate filamentous fungi is of great value for improving the low yields of bioactive metabolites and expanding chemical diversity. The CRISPR-Cas9-based system has become a common platform for genome editing in a variety of organisms. However, recent application of this technology in filamentous fungi is limited to model strains, a versatile method for efficient gene disruption in different fungi is lacking. Here, we investigated the utility of the CRISPR-Cas9 system in a less-studied fungus Nodulisporium sp. (No. 65-12-7-1), and we have developed an efficient CRISPR-Cas9-based gene disruption strategy by simultaneous transformation of in vitro transcriptional gRNA and the linear maker gene cassette into the Cas9-expressing fungi. We found that the linear marker gene cassette could not only allow for selection of transformants, but also significantly enhance the gene disruption efficiency by inserting itself into the Cas9 cut site. Moreover, the above approach also demonstrated its efficiency in two other phylogenetically distinct strains Aspergillus oryzae NSAR1 and Sporormiella minima (No. 40-1-4-1) from two different classes of Ascomycota. These results suggested that a versatile CRISPR-Cas9-based gene disruption method in filamentous fungi was established.Yan-Mei ZhengFu-Long LinHao GaoGen ZouJiang-Wei ZhangGao-Qian WangGuo-Dong ChenZhi-Hua ZhouXin-Sheng YaoDan HuNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 7, Iss 1, Pp 1-10 (2017)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Yan-Mei Zheng Fu-Long Lin Hao Gao Gen Zou Jiang-Wei Zhang Gao-Qian Wang Guo-Dong Chen Zhi-Hua Zhou Xin-Sheng Yao Dan Hu Development of a versatile and conventional technique for gene disruption in filamentous fungi based on CRISPR-Cas9 technology
description	Abstract Filamentous fungi represent an invaluable source of pharmaceutically active compounds. The development of versatile methods to genetically manipulate filamentous fungi is of great value for improving the low yields of bioactive metabolites and expanding chemical diversity. The CRISPR-Cas9-based system has become a common platform for genome editing in a variety of organisms. However, recent application of this technology in filamentous fungi is limited to model strains, a versatile method for efficient gene disruption in different fungi is lacking. Here, we investigated the utility of the CRISPR-Cas9 system in a less-studied fungus Nodulisporium sp. (No. 65-12-7-1), and we have developed an efficient CRISPR-Cas9-based gene disruption strategy by simultaneous transformation of in vitro transcriptional gRNA and the linear maker gene cassette into the Cas9-expressing fungi. We found that the linear marker gene cassette could not only allow for selection of transformants, but also significantly enhance the gene disruption efficiency by inserting itself into the Cas9 cut site. Moreover, the above approach also demonstrated its efficiency in two other phylogenetically distinct strains Aspergillus oryzae NSAR1 and Sporormiella minima (No. 40-1-4-1) from two different classes of Ascomycota. These results suggested that a versatile CRISPR-Cas9-based gene disruption method in filamentous fungi was established.
format	article
author	Yan-Mei Zheng Fu-Long Lin Hao Gao Gen Zou Jiang-Wei Zhang Gao-Qian Wang Guo-Dong Chen Zhi-Hua Zhou Xin-Sheng Yao Dan Hu
author_facet	Yan-Mei Zheng Fu-Long Lin Hao Gao Gen Zou Jiang-Wei Zhang Gao-Qian Wang Guo-Dong Chen Zhi-Hua Zhou Xin-Sheng Yao Dan Hu
author_sort	Yan-Mei Zheng
title	Development of a versatile and conventional technique for gene disruption in filamentous fungi based on CRISPR-Cas9 technology
title_short	Development of a versatile and conventional technique for gene disruption in filamentous fungi based on CRISPR-Cas9 technology
title_full	Development of a versatile and conventional technique for gene disruption in filamentous fungi based on CRISPR-Cas9 technology
title_fullStr	Development of a versatile and conventional technique for gene disruption in filamentous fungi based on CRISPR-Cas9 technology
title_full_unstemmed	Development of a versatile and conventional technique for gene disruption in filamentous fungi based on CRISPR-Cas9 technology
title_sort	development of a versatile and conventional technique for gene disruption in filamentous fungi based on crispr-cas9 technology
publisher	Nature Portfolio
publishDate	2017
url	https://doaj.org/article/435d45484750440791e74b5b38caee94
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Development of a versatile and conventional technique for gene disruption in filamentous fungi based on CRISPR-Cas9 technology

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