title: Gene targeting in the oil-producing fungus Mortierella alpina 1S-4 and construction of a strain producing a valuable polyunsaturated fatty acid.
abstract: To develop an efficient gene-targeting system in Mortierella alpina 1S-4, we identified the ku80 gene encoding the Ku80 protein, which is involved in the nonhomologous end-joining pathway in genomic double-strand break (DSB) repair, and constructed ku80 gene-disrupted strains via single-crossover homologous recombination. The Δku80 strain from M. alpina 1S-4 showed no negative effects on vegetative growth, formation of spores, and fatty acid productivity, and exhibited high sensitivity to methyl methanesulfonate, which causes DSBs. Dihomo-γ-linolenic acid (DGLA)-producing strains were constructed by disruption of the Δ5-desaturase gene, encoding a key enzyme of bioconversion of DGLA to ARA, using the Δku80 strain as a host strain. The significant improvement of gene-targeting efficiency was not observed by disruption of the ku80 gene, but the construction of DGLA-producing strain by disruption of the Δ5-desaturase gene was succeeded using the Δku80 strain as a host strain. This report describes the first study on the identification and disruption of the ku80 gene in zygomycetes and construction of a DGLA-producing transformant using a gene-targeting system in M. alpina 1S-4.
authors.affiliation: Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-oiwakecho, Sakyo-ku, Kyoto, 606-8502, Japan.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Kikukawa
authors.affiliation: Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-oiwakecho, Sakyo-ku, Kyoto, 606-8502, Japan.|Institute of Technology and Science, Tokushima University, 2-1 Minami-josanjima, Tokushima, 770-8506, Japan.
authors.forename: Eiji
authors.initials: E
authors.lastname: Sakuradani
authors.affiliation: Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-oiwakecho, Sakyo-ku, Kyoto, 606-8502, Japan.
authors.forename: Masato
authors.initials: M
authors.lastname: Nakatani
authors.affiliation: Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-oiwakecho, Sakyo-ku, Kyoto, 606-8502, Japan.|Research Unit for Physiological Chemistry, Kyoto University, Kitashirakawa-oiwakecho, Sakyo-ku, Kyoto, 606-8502, Japan.
authors.forename: Akinori
authors.initials: A
authors.lastname: Ando
authors.affiliation: Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-oiwakecho, Sakyo-ku, Kyoto, 606-8502, Japan.
authors.forename: Tomoyo
authors.initials: T
authors.lastname: Okuda
authors.affiliation: Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-oiwakecho, Sakyo-ku, Kyoto, 606-8502, Japan.
authors.forename: Takaiku
authors.initials: T
authors.lastname: Sakamoto
authors.affiliation: Research Institute, Suntory Global Innovation Center Ltd., 1-1-1 Wakayamadai, Shimamoto-cho, Mishima-gun, Osaka, 618-8503, Japan.
authors.forename: Misa
authors.initials: M
authors.lastname: Ochiai
authors.affiliation: Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-oiwakecho, Sakyo-ku, Kyoto, 606-8502, Japan.|Department of Bioscience and Biotechnology, Faculty of Bioenvironmental Science, Kyoto Gakuen University, 1-1 Nanjo, Sogabe, Kameoka, 621-8555, Japan.
authors.forename: Sakayu
authors.initials: S
authors.lastname: Shimizu
authors.affiliation: Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-oiwakecho, Sakyo-ku, Kyoto, 606-8502, Japan. ogawa@kais.kyoto-u.ac.jp.|Research Unit for Physiological Chemistry, Kyoto University, Kitashirakawa-oiwakecho, Sakyo-ku, Kyoto, 606-8502, Japan. ogawa@kais.kyoto-u.ac.jp.
authors.forename: Jun
authors.initials: J
authors.lastname: Ogawa
mesh_terms: D015126:8,11,14-Eicosatrienoic Acid / Q000378:metabolism*; D016718:Arachidonic Acid / Q000378:metabolism; D004247:DNA / Q000235:genetics* / Q000378:metabolism; D053903:DNA Breaks, Double-Stranded; D059766:DNA End-Joining Repair; D004271:DNA, Fungal / Q000235:genetics* / Q000378:metabolism; D000090943:Delta-5 Fatty Acid Desaturase; D044943:Fatty Acid Desaturases / Q000172:deficiency / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism; D018390:Gene Targeting; D005818:Genetic Engineering; D059765:Homologous Recombination; D008698:Mesylates / Q000494:pharmacology; D020094:Mortierella / Q000145:classification / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D010802:Phylogeny
chemical_list: D004271:DNA, Fungal; D000090943:Delta-5 Fatty Acid Desaturase; D005656:Fungal Proteins; D008698:Mesylates; C045880:methanesulfonic acid; D016718:Arachidonic Acid; D004247:DNA; D044943:Fatty Acid Desaturases; D015126:8,11,14-Eicosatrienoic Acid
keywords: Dihomo-γ-linolenic acid; Gene targeting; Homologous recombination; Ku80; Mortierella alpina; Δ5-desaturase