title: An efficient blue-white screening based gene inactivation system for Streptomyces.
abstract: 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.
authors: State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, NO.1 Beichen West Road, Chaoyang District, Beijing, 100101, China.
authors: Pengwei
authors: P
authors: Li
authors: Jine
authors: J
authors: Li
authors: Zhengyan
authors: Z
authors: Guo
authors: Wei
authors: W
authors: Tang
authors: Jianshan
authors: J
authors: Han
authors: Xiangxi
authors: X
authors: Meng
authors: Tingting
authors: T
authors: Hao
authors: Yaxin
authors: Y
authors: Zhu
authors: Lixin
authors: L
authors: Zhang
authors: Yihua
authors: Y
authors: Chen
mesh_terms: D003433:Crosses, Genetic; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D020868:Gene Silencing; D005798:Genes, Bacterial; D017930:Genes, Reporter; D005820:Genetic Testing / Q000379:methods*; D005822:Genetic Vectors; D005827:Genetics, Microbial / Q000379:methods*; D008967:Molecular Biology / Q000379:methods*; D008969:Molecular Sequence Data; D010881:Piperidones / Q000378:metabolism; D010957:Plasmids; D011995:Recombination, Genetic; D017422:Sequence Analysis, DNA; D013302:Streptomyces / Q000235:genetics* / Q000302:isolation & purification*
chemical_list: D004269:DNA, Bacterial; D010881:Piperidones; C446045:indigoidine