Genome engineering and direct cloning of antibiotic gene clusters via phage ϕBT1 integrase-mediated site-specific recombination in Streptomyces.
Several strategies have been used to clone large DNA fragments directly from bacterial genome. Most of these approaches are based on different site-specific recombination systems consisting of a specialized recombinase and its target sites. In this study, a novel strategy based on phage ϕBT1 integrase-mediated site-specific recombination was developed, and used for simultaneous Streptomyces genome engineering and cloning of antibiotic gene clusters. This method has been proved successful for the cloning of actinorhodin gene cluster from Streptomyces coelicolor M145, napsamycin gene cluster and daptomycin gene cluster from Streptomyces roseosporus NRRL 15998 at a frequency higher than 80%. Furthermore, the system could be used to increase the titer of antibiotics as we demonstrated with actinorhodin and daptomycin, and it will be broadly applicable in many Streptomyces.
1] State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China [2] University of Chinese Academy of Sciences, Beijing, China.
Deyao
D
Du
1] State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China [2] Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, College of Biotechnology, Tianjin University of Science and Technology, Tianjin, China.
Lu
L
Wang
State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China.
Yuqing
Y
Tian
Key Laboratory of Industrial Fermentation Microbiology, Ministry of Education, College of Biotechnology, Tianjin University of Science and Technology, Tianjin, China.
Hao
H
Liu
State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China.
Huarong
H
Tan
State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing, China.
Guoqing
G
Niu
D000880:Anthraquinones / Q000378:metabolism; D000900:Anti-Bacterial Agents / Q000096:biosynthesis*; D023181:Antimicrobial Cationic Peptides; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001435:Bacteriophages / Q000201:enzymology / Q000235:genetics; D001665:Binding Sites / Q000235:genetics; D003001:Cloning, Molecular / Q000379:methods; D017576:Daptomycin / Q000096:biosynthesis; D016680:Genome, Bacterial / Q000235:genetics; D019426:Integrases / Q000235:genetics* / Q000378:metabolism; D060847:Metabolic Engineering / Q000379:methods*; D008957:Models, Genetic; D005810:Multigene Family / Q000235:genetics*; D010455:Peptides / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011995:Recombination, Genetic / Q000235:genetics*; D015203:Reproducibility of Results; D013302:Streptomyces / Q000235:genetics* / Q000378:metabolism; D048372:Streptomyces coelicolor / Q000235:genetics; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism; D000095622:Benzoisochromanequinones
D000880:Anthraquinones; D000900:Anti-Bacterial Agents; D023181:Antimicrobial Cationic Peptides; D001426:Bacterial Proteins; D017576:Daptomycin; D019426:Integrases; D010455:Peptides; D014764:Viral Proteins; C088538:napsamycin D; C013390:actinorhodin; D000095622:Benzoisochromanequinones