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Gene targeting in the oil-producing fungus Mortierella alpina 1S-4 and construction of a strain producing a valuable polyunsaturated fatty acid

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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.

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Acknowledgments

This work was partially supported by Grants-in Aid for Scientific Research of Japan (Numbers 22380051 to E. Sakuradani and 23248014 to J. Ogawa), the Program for Promotion of Basic and Applied Researches for Innovations in Bio-oriented Industry (BRAIN) of Japan, and the Advanced Low Carbon Technology Research and Development Program (ALCA) of Japan.

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Author notes

  1. Jun Ogawa

    Present address: Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-oiwakecho, Sakyo-ku, Kyoto, 606-8502, Japan

Authors and Affiliations

  1. Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Kitashirakawa-oiwakecho, Sakyo-ku, Kyoto, 606-8502, Japan

    Hiroshi Kikukawa, Eiji Sakuradani, Masato Nakatani, Akinori Ando, Tomoyo Okuda, Takaiku Sakamoto & Sakayu Shimizu

  2. Institute of Technology and Science, Tokushima University, 2-1 Minami-josanjima, Tokushima, 770-8506, Japan

    Eiji Sakuradani

  3. Research Unit for Physiological Chemistry, Kyoto University, Kitashirakawa-oiwakecho, Sakyo-ku, Kyoto, 606-8502, Japan

    Akinori Ando & Jun Ogawa

  4. Research Institute, Suntory Global Innovation Center Ltd., 1-1-1 Wakayamadai, Shimamoto-cho, Mishima-gun, Osaka, 618-8503, Japan

    Misa Ochiai

  5. Department of Bioscience and Biotechnology, Faculty of Bioenvironmental Science, Kyoto Gakuen University, 1-1 Nanjo, Sogabe, Kameoka, 621-8555, Japan

    Sakayu Shimizu

Authors
  1. Hiroshi Kikukawa
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  2. Eiji Sakuradani
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Correspondence to Jun Ogawa.

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Communicated by G. Goldman.

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Kikukawa, H., Sakuradani, E., Nakatani, M. et al. Gene targeting in the oil-producing fungus Mortierella alpina 1S-4 and construction of a strain producing a valuable polyunsaturated fatty acid. Curr Genet 61, 579–589 (2015). https://doi.org/10.1007/s00294-015-0481-2

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