Development of a CRISPR/Cpf1 system for multiplex gene editing in Aspergillus oryzae

文献类型： 外文期刊

第一作者： Chen, Tianming

作者： Chen, Tianming;Chen, Ziming;Li, Yuzhen;Yao, Lihua;Zeng, Bin;Zhang, Zhe;Zhang, Huanxin;Zeng, Bin

作者机构：

关键词： CRISPR; Cpf1 system; Multiplex gene editing; Filamentous fungi; Aspergillus oryzae

期刊名称：FOLIA MICROBIOLOGICA （ 影响因子：2.6； 五年影响因子：2.6 ）

ISSN： 0015-5632

年卷期： 2023 年

页码：

收录情况： SCI

摘要： CRISPR/Cas technology is a powerful tool for genome engineering in Aspergillus oryzae as an industrially important filamentous fungus. Previous study has reported the application of the CRISPR/Cpf1 system based on the Cpf1 (LbCpf1) from Lachnospiraceae bacterium in A. oryzae. However, multiplex gene editing have not been investigated using this system. Here, we presented a new CRISPR/Cpf1 multiplex gene editing system in A. oryzae, which contains the Cpf1 nuclease (FnCpf1) from Francisella tularensis subsp. novicida U112 and CRISPR-RNA expression cassette. The crRNA cassette consisted of direct repeats and guide sequences driven by the A. oryzae U6 promoter and U6 terminator. Using the constructed FnCpf1 gene editing system, the wA and pyrG genes were mutated successfully. Furthermore, simultaneous editing of wA and pyrG genes in A. oryzae was performed using two guide sequences targeting these gene loci in a single crRNA array. This promising CRISPR/Cpf1 genome-editing system provides a powerful tool for genetically engineering A. oryzae.

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