Construction of linear plasmid vector for cloning of giant gene cluster for secondary metabolite biosynthesis

2015 Fiscal Year Final Research Report

• Project/Area Number: 25450112
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Applied microbiology
• Research Institution: Kitasato University
• Principal Investigator: KOMATSU MAMORU 北里大学, 感染制御科学府, 講師 (40414057)
• Project Period (FY): 2013-04-01 – 2016-03-31
• Keywords: 二次代謝産物生産 / 異種発現 / 接合伝達 / 放線菌 / 生合成遺伝子 / 線状プラスミド

Outline of Final Research Achievements

We have constructed the large-deletion mutants of S. avermitilis (SUKA) for expression of exogenous gene clusters for secondary metabolite biosynthesis. The entire biosynthetic gene cluster for polyketide compound is quite large in comparison with biosynthetic gene clusters for other metabolites and some of them are ~ 100 kb. In order to achieve the heterologous expression of these giant biosynthetic gene clusters in SUKA, more efficient methods for gene introduction are indispensable. Most Streptomyces microorganisms strongly restrict DNA prepared in E. coli because they possess unique methyl-specific restriction system. However, S. lividans is not an ideal host since it possesses a weak methyl-specific restriction system. In this study, we have developed efficient heterologous expression system. Procedures are based on integrating BAC vector, a preferable transformation host (S. lividans) and transfer of linear plasmid vector by interspecific mating between S. lividans and SUKA.

Free Research Field

応用微生物