title: Disruption of lig4 improves gene targeting efficiency in the oleaginous fungus Mortierella alpina 1S-4.
abstract: The oil-producing zygomycete Mortierella alpina 1S-4 is known to accumulate beneficial polyunsaturated fatty acids. We identified the lig4 gene that encodes for a DNA ligase 4 homolog, which functions to repair double strand breaks by non-homologous end joining. We disrupted the lig4 gene to improve the gene targeting efficiency in M. alpina. The M. alpina 1S-4 Δlig4 strains showed no defect in vegetative growth, formation of spores, and fatty acid production, but exhibited high sensitivity to methyl methansulfonate, an agent that causes DNA double-strand breaks. Importantly, gene replacement of ura5 marker by CBXB marker occurred in 67% of Δlig4 strains and the gene targeting efficiency was 21-fold greater than that observed in disruption of the lig4 gene in the M. alpina 1S-4 host strain. Further metabolic engineering of the Δlig4 strains is expected to result in strains that produce higher levels of rare and beneficial polyunsaturated fatty acids and contribute to basic research on the zygomycete.
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; 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: 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; Research Unit for Physiological Chemistry, Kyoto University, Kitashirakawa-Oiwakecho, Sakyo-ku, Kyoto 606-8502, Japan. Electronic address: ogawa@kais.kyoto-u.ac.jp.
authors.forename: Jun
authors.initials: J
authors.lastname: Ogawa
mesh_terms: D011088:DNA Ligases / Q000235:genetics* / Q000378:metabolism; D005656:Fungal Proteins / Q000235:genetics* / Q000378:metabolism; D055785:Gene Knockdown Techniques; D018390:Gene Targeting / Q000379:methods*; D020094:Mortierella / Q000201:enzymology / Q000235:genetics*
chemical_list: D005656:Fungal Proteins; D011088:DNA Ligases
keywords: DNA ligase 4 (lig4); Gene targeting; Homologous recombination