ku70 and ku80 null mutants improve the gene targeting frequency in Monascus ruber M7

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• 作者：Yi He (3)
  Qingpei Liu (3)
  Yanchun Shao (1) (3)
  Fusheng Chen (1) (2) (3)
  
• 关键词：Monascus ruber ; Gene replacement frequency ; ku70 and ku80 ; Non ; homologous end ; joining ; Homologous recombination
• 刊名：Applied Microbiology and Biotechnology
• 出版年：2013
• 出版时间：June 2013
• 年：2013
• 卷：97
• 期：11
• 页码：4965-4976
• 全文大小：607KB
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• 作者单位：Yi He (3)
  Qingpei Liu (3)
  Yanchun Shao (1) (3)
  Fusheng Chen (1) (2) (3)
  
  3. College of Food Science and Technology, Huazhong Agricultural University, Wuhan, 430070, Hubei Province, People’s Republic of China
  1. National Key Laboratory of Agro-Microbiology, Huazhong Agricultural University, Wuhan, 430070, Hubei Province, People’s Republic of China
  2. Key Laboratory of Environment Correlative Dietology, Huazhong Agricultural University, Ministry of Education, Wuhan, 430070, Hubei Province, People’s Republic of China
  
• ISSN：1432-0614

文摘

Normally, gene targeting by homologous recombination occurs rarely during a transformation process since non-homologous recombination is predominant in filamentous fungi. In our previous researches, the average gene replacement frequency (GRF) in Monascus ruber M7 was as low as 15?%. To develop a highly efficient gene targeting system for M. ruber M7, two M. ruber M7 null mutants of ku70 (MrΔku70) and ku80 (MrΔku80) were constructed which had no apparent defects in the development including vegetative growth, colony phenotype, microscopic morphology and spore yield compared with M. ruber M7. In addition, the production of some significant secondary metabolites such as pigments and citrinin had no differences between the two disruptants and the wild-type strain. Further results revealed that the GRFs of triA (encoding a putative acetyltransferase) were 42.2?% and 61.5?% in the MrΔku70 and MrΔku80 strains, respectively, while it was only about 20?% in M. ruber M7. Furthermore, GRFs of these two disruptants at other loci (the pigE, fmdS genes in MrΔku70 and the ku70 gene in MrΔku80) were investigated, and the results indicated that GRFs in the MrΔku70 strain and the MrΔku80 strain were doubled and tripled compared with that in M. ruber M7, respectively. Therefore, the ku70 and ku80 null mutants of M. ruber M7, especially the ku80-deleted strain, will be excellent hosts for efficient gene targeting.