title: Recombinational cloning of the antibiotic biosynthetic gene clusters in linear plasmid SCP1 of Streptomyces coelicolor A3(2).
abstract: The model organism Streptomyces coelicolor A3(2) harbors a 356-kb linear plasmid, SCP1. We report here development of a recombinational cloning method for deleting large segment from one telomere of SCP1 followed by replacing with the telomere of pSLA2 and sequentially inserting with the overlapping cosmids in vivo. The procedure depends on homologous recombination coupled with cleavage at telomere termini by telomere terminal protein. Using this procedure, we cloned the 81-kb avermectin and the 76-kb spinosad biosynthetic gene clusters into SCP1. Heterologous expression of avermectin production in S. coelicolor was detected. These results demonstrate the utility of SCP1 for cloning large DNA segments such as antibiotic biosynthetic gene clusters.
authors.affiliation: Key laboratory of Synthetic Biology, Shanghai Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, the Chinese Academy of Sciences, Shanghai, China.
authors.forename: Ran
authors.initials: R
authors.lastname: Zhang
authors.forename: Haiyang
authors.initials: H
authors.lastname: Xia
authors.forename: Qingyu
authors.initials: Q
authors.lastname: Xu
authors.forename: Fujun
authors.initials: F
authors.lastname: Dang
authors.forename: Zhongjun
authors.initials: Z
authors.lastname: Qin
mesh_terms: D000900:Anti-Bacterial Agents / Q000096:biosynthesis*; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular / Q000379:methods*; D059765:Homologous Recombination; D005810:Multigene Family; D010957:Plasmids / Q000235:genetics* / Q000378:metabolism; D048372:Streptomyces coelicolor / Q000235:genetics* / Q000378:metabolism
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins
keywords: Streptomyces; linear plasmid; recombinational cloning