An efficient blue-white screening based gene inactivation system for Streptomyces

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• 作者：Pengwei Li ; Jine Li ; Zhengyan Guo ; Wei Tang…
• 关键词：Streptomyces ; Indigoidine ; Gene inactivation ; Blue ; white screening
• 刊名：Applied Microbiology and Biotechnology
• 出版年：2015
• 出版时间：February 2015
• 年：2015
• 卷：99
• 期：4
• 页码：1923-1933
• 全文大小：1,617 KB
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• 作者单位：Pengwei Li (1) (3)
  Jine Li (1)
  Zhengyan Guo (1)
  Wei Tang (1) (3)
  Jianshan Han (1) (3)
  Xiangxi Meng (1) (3)
  Tingting Hao (1) (3)
  Yaxin Zhu (1)
  Lixin Zhang (2)
  Yihua Chen (1)
  
  1. State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, NO.1 Beichen West Road, Chaoyang District, Beijing, 100101, China
  3. University of Chinese Academy of Sciences, Beijing, 100049, China
  2. CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of Microbiology, Chinese Academy of Sciences, Beijing, 100101, China
  
• 刊物类别：Chemistry and Materials Science
• 刊物主题：Chemistry
  Biotechnology
  Microbiology
  Microbial Genetics and Genomics
  
• 出版者：Springer Berlin / Heidelberg
• ISSN：1432-0614

文摘

Streptomyces is studied intensively for its outstanding ability to produce bioactive secondary metabolites and for its complicated morphological differentiation process. A classical genetic manipulation system for Streptomyces has been developed and widely used in the community for a long time, using antibiotic resistance markers to select for double-crossover mutants. The screening process is always laborious and time-consuming. However, the lack of a suitable chromogenic reporter for Streptomyces has limited the use of color-based screening system to simplify the selection process for double-crossover mutants. In this study, a blue reporter system for Streptomyces has been established by mining an indigoidine synthetase gene (idgS) from Streptomyces lavendulae CGMCC 4.1386, leading to the development of a time-saving gene inactivation system for Streptomyces by simple blue-white screening. A series of Streptomyces suicide and temperature-sensitive plasmids containing the idgS reporter cassette were constructed and used successfully to inactivate genes in Streptomyces, allowing a simple and efficient screening method to differentiate the colonies for double-crossover (white) and single-crossover (blue) mutants. Inactivation of the putative γ-butyrolactone synthase gene afsA-y via the idgS-based blue-white screening method revealed that the paulomycin production is negatively controlled by afsA-y in Streptomyces sp. YN86.