Rapid Reconstitution of Biosynthetic Machinery for Fungal Metabolites in Aspergillus oryzae: Total Biosynthesis of Aflatrem |
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| Authors: | Koichi Tagami Dr Atsushi Minami Ryuya Fujii Dr Chengwei Liu Dr Mizuki Tanaka Prof Dr Katsuya Gomi Prof Dr Tohru Dairi Prof Dr Hideaki Oikawa |
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| Affiliation: | 1. Division of Chemistry, Graduate School of Science, Hokkaido University, Sapporo 060‐0810 (Japan);2. Graduate School of Agricultural Science, Tohoku University, Sendai 981‐8555 (Japan);3. Graduate School of Engineering, Hokkaido University, Sapporo 060‐8628 (Japan) |
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| Abstract: | Reconstitution of the biosynthetic machinery for fungal secondary metabolites in Aspergillus oryzae provides an opportunity both for stepwise determination of the biosynthetic pathways and the total biosynthesis of fungal natural products. However, to maximize the utility of the reconstitution system, a simple and rapid strategy for the introduction of heterologous genes into A. oryzae is required. In this study, we demonstrated an effective method for introducing multiple genes involved in the biosynthesis of fungal metabolites by using the expression vectors pUARA2 and pUSA2, each of which contains two cloning sites. The successful introduction of all the aflatrem biosynthetic genes (seven genes in total) after two rounds of transformation enabled the total biosynthesis of aflatrem. This rapid reconstitution strategy will facilitate the functional analysis of the biosynthetic machinery of fungal metabolites. |
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| Keywords: | aflatrem Aspergillus oryzae gene expression indole diterpene total biosynthesis |
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