Development of a Synthetic Oxytetracycline-Inducible Expression System for Streptomycetes Using de Novo Characterized Genetic Parts

详细信息    查看全文

• 作者：Weishan Wang ; Tongjian Yang ; Yihong Li ; Shanshan Li ; Shouliang Yin ; Kathryn Styles ; Christophe Corre ; Keqian Yang
• 刊名：ACS Synthetic Biology
• 出版年：2016
• 出版时间：July 15, 2016
• 年：2016
• 卷：5
• 期：7
• 页码：765-773
• 全文大小：532K
• 年卷期：0
• ISSN：2161-5063

文摘

Precise control of gene expression using exogenous factors is of great significance. To develop ideal inducible expression systems for streptomycetes, new genetic parts, oxytetracycline responsive repressor OtrR, operator otrO, and promoter otrBp from Streptomyces rimosus, were selected de novo and characterized in vivo and in vitro. OtrR showed strong affinity to otrO (K_D = 1.7 × 10^–10 M) and oxytetracycline induced dissociation of the OtrR/DNA complex in a concentration-dependent manner. On the basis of these genetic parts, a synthetic inducible expression system Potr* was optimized. Induction of Potr* with 0.01–4 μM of oxytetracycline triggered a wide-range expression level of gfp reporter gene in different Streptomyces species. Benchmarking Potr* against the widely used constitutive promoters ermE* and kasOp* revealed greatly enhanced levels of expression when Potr* was fully induced. Finally, Potr* was used as a tool to activate and optimize the expression of the silent jadomycin biosynthetic gene cluster in Streptomyces venezuelae. Altogether, the synthetic Potr* presents a new versatile tool for fine-tuning gene expression in streptomycetes.