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Rapid Reconstitution of Biosynthetic Machinery for Fungal Metabolites in Aspergillus oryzae : Total Biosynthesis of Aflatrem
Author(s) -
Tagami Koichi,
Minami Atsushi,
Fujii Ryuya,
Liu Chengwei,
Tanaka Mizuki,
Gomi Katsuya,
Dairi Tohru,
Oikawa Hideaki
Publication year - 2014
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.201402195
Subject(s) - aspergillus oryzae , biosynthesis , biology , gene , cloning (programming) , heterologous , biochemistry , heterologous expression , secondary metabolism , aspergillus , metabolic engineering , transformation (genetics) , enzyme , microbiology and biotechnology , recombinant dna , computer science , programming language
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.