Recombinational cloning of the antibiotic biosynthetic gene clusters in linear plasmid SCP1 of Streptomyces coelicolor A3(2)

Authors

  • Ran Zhang,

    1. Key laboratory of Synthetic Biology, Shanghai Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, the Chinese Academy of Sciences, Shanghai, China
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  • Haiyang Xia,

    1. Key laboratory of Synthetic Biology, Shanghai Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, the Chinese Academy of Sciences, Shanghai, China
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  • Qingyu Xu,

    1. Key laboratory of Synthetic Biology, Shanghai Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, the Chinese Academy of Sciences, Shanghai, China
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  • Fujun Dang,

    1. Key laboratory of Synthetic Biology, Shanghai Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, the Chinese Academy of Sciences, Shanghai, China
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  • Zhongjun Qin

    Corresponding author
    • Key laboratory of Synthetic Biology, Shanghai Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, the Chinese Academy of Sciences, Shanghai, China
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Correspondence: Zhongjun Qin, Shanghai Institute of Plant Physiology and Ecology, 300 Fenglin Road, Shanghai 200032, China. Tel.: +86 21 54924171; fax: +86 21 54924176; e-mail: qin@sibs.ac.cn

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.

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