title: Establishment of an overall transformation system for an oil-producing filamentous fungus, Mortierella alpina 1S-4.
abstract: Oil-producing fungus Mortierella alpina 1S-4 is an industrial strain. To determine its physiological properties and to clarify the biosynthetic pathways for polyunsaturated fatty acids, a transformation system for this fungus was established using a derivative of it, i.e., a ura5- mutant lacking orotate phosphoribosyl transferase (OPRTase, EC.2.4.2.10) activity. Transformation with a vector containing the homologous ura5 gene as a marker was successfully performed using microprojectile bombardment, other methods frequently used for transformation, such as the protoplasting, lithium acetate, or electroporation methods, not giving satisfactory results. As a result, two types of transformants were obtained: a few stable transformants overexpressing the ura5 gene, and many unstable transformants showing OPRTase activity comparable to that of the wild-type strain. The results of quantitative PCR indicated that the stable transformants could retain the ura5 genes originating from the transformation vector regardless of the culture conditions. On the other hand, unstable transformants easily lost the marker gene under uracil-containing conditions, as expected. In this paper, we report that an overall transformation system for this fungus was successfully established, and propose how to select useful transformants as experimental and industrial strains.
authors.affiliation: Laboratory of Fermentation Physiology and Applied Microbiology, Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, 606-8502, Kyoto, Japan.
authors.forename: Seiki
authors.initials: S
authors.lastname: Takeno
authors.forename: Eiji
authors.initials: E
authors.lastname: Sakuradani
authors.forename: Shoichi
authors.initials: S
authors.lastname: Murata
authors.forename: Misa
authors.initials: M
authors.lastname: Inohara-Ochiai
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Kawashima
authors.forename: Toshihiko
authors.initials: T
authors.lastname: Ashikari
authors.forename: Sakayu
authors.initials: S
authors.lastname: Shimizu
mesh_terms: D000085:Acetates / Q000494:pharmacology; D003470:Culture Media / Q000737:chemistry; D018274:Electroporation; D005800:Genes, Fungal; D005822:Genetic Vectors; D020094:Mortierella / Q000235:genetics* / Q000378:metabolism; D009154:Mutation; D009821:Oils / Q000378:metabolism; D009962:Orotate Phosphoribosyltransferase / Q000235:genetics* / Q000378:metabolism; D011523:Protoplasts; D012641:Selection, Genetic; D014170:Transformation, Genetic
chemical_list: D000085:Acetates; D003470:Culture Media; D009821:Oils; D009962:Orotate Phosphoribosyltransferase; C488804:lithium acetate