Construction of single, double, or triple mutants within kojic acid synthesis genes kojA, kojR, and kojT by the CRISPR/Cas9 tool in Aspergillus oryzae

Author(s):  
Yuzhen Li ◽  
Huanxin Zhang ◽  
Ziming Chen ◽  
Junxia Fan ◽  
Tianming Chen ◽  
...  
2011 ◽  
Vol 75 (9) ◽  
pp. 1832-1834 ◽  
Author(s):  
Ken ODA ◽  
Akiko KOBAYASHI ◽  
Shinichi OHASHI ◽  
Motoaki SANO

2021 ◽  
Vol 204 (1) ◽  
Author(s):  
Yuzhen Li ◽  
Huanxin Zhang ◽  
Ziming Chen ◽  
Junxia Fan ◽  
Tianming Chen ◽  
...  

2021 ◽  
Author(s):  
Junxia Fan ◽  
Huanxin Zhang ◽  
Yuzhen Li ◽  
Ziming Chen ◽  
Tianming Chen ◽  
...  

Abstract The CRISPR/Cas9 system has become a great tool for target gene knock-out in filamentous fungi. It is laborious and time-consuming that identification mutants from a large number of transformants through PCR or enzyme-cut method. Here, we first developed a CRISPR/Cas9 system in Aspergillus oryzae using AMA1-based autonomously replicating plasmid and Cas9 under the control of the Aspergillus nidulans gpdA promoter. By the genome editing technique, we successfully obtained mutations within each target gene in Aspergillus oryzae. Then, we put the protospacer sequence of a target gene and its protospacer adjacent motif (PAM) behind the start codon “ATG” of DsRed, yielding the non‑functional DsRed (nDsRed) reporter gene, and the nDsRed reporter gene could be rescued after successful targeted editing. Moreover, this method was also applied by targeting the kojic acid synthesis gene kojA, and the transformants with DsRed activity were found to harbor targeted mutations in kojA. These results suggest that the nDsRed can be used as a powerful tool to facilitate the identification of mutants generated by CRISPR/Cas9 in Aspergillus oryzae.


2021 ◽  
Author(s):  
Yuzhen Li ◽  
Huanxin Zhang ◽  
Ziming Chen ◽  
Junxia Fan ◽  
Tianming Chen ◽  
...  

Abstract Aspergillus oryzae usually suffers from oxidative stress during the process of aerobic fermentation. However, there is little information on what genes involve in oxidative stress of A. oryzae. Here, we found that the expression of a novel gene Aohps1 was induced during the growth and development of A. oryzae with and without oxidative stress. Sequence analysis revealed that Aohps1 has four transmembrane regions and is conserved in Aspergillus species. Overexpression and deletion of Aohpi1 caused the growth defects with and without oxidative stress, including mycelium growth, conidia formation and biomass. Meanwhile, the Aohpi1 overexpression strain exhibited more sensitivity to oxidative stress than the Aohpi1 disrupted mutant. Furthermore, overexpression and disruption of Aohps1 resulted in the inhibition of kojic acid production with and without oxidative stress, consistent with the reduced expression of kojA that directly contributed to the synthesis of kojic acid. Additionally, the yield of kojic acid is less in the Aohps1 overexpression strain than in the Aohps1 deletion mutant under oxidative stress. Collectively, the discovery of Aohps1 provides new insights into oxidative stress and kojic acid synthesis in A. oryzae.


Author(s):  
Yuzhen Li ◽  
Huanxin Zhang ◽  
Ziming Chen ◽  
Junxia Fan ◽  
Tianming Chen ◽  
...  

2011 ◽  
Vol 11 (1) ◽  
pp. 263-267 ◽  
Author(s):  
Fabio C. Chaves ◽  
Thomas J. Gianfagna ◽  
Madhu Aneja ◽  
Francisco Posada ◽  
Stephen W. Peterson ◽  
...  

2001 ◽  
Vol 91 (3) ◽  
pp. 272-276 ◽  
Author(s):  
Takafumi Futamura ◽  
Mitsuyasu Okabe ◽  
Takayoshi Tamura ◽  
Kyouko Toda ◽  
Toshio Matsunobu ◽  
...  

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