Mycobiology

  • 제49권5호
  • /
  • Pages.491-497
  • /
  • 2021
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  • 1229-8093(pISSN)
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  • 2092-9323(eISSN)
한국균학회 (The Korean Society of Mycology)
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Optimization of Agrobacterium tumefaciens-Mediated Transformation of Xylaria grammica EL000614, an Endolichenic Fungus Producing Grammicin

  • Jeong, Min-Hye (Department of Plant Medicine, Sunchon National University) ;
  • Kim, Jung A. (Microorganism Resources Division, National Institute of Biological Resources) ;
  • Kang, Seogchan (Department of Plant Pathology & Environmental Microbiology, The Pennsylvania State University) ;
  • Choi, Eu Ddeum (Department of Plant Medicine, Sunchon National University) ;
  • Kim, Youngmin (Department of Plant Medicine, Sunchon National University) ;
  • Lee, Yerim (Department of Plant Medicine, Sunchon National University) ;
  • Jeon, Mi Jin (Microorganism Resources Division, National Institute of Biological Resources) ;
  • Yu, Nan Hee (Department of Agricultural Chemistry, Institute of Environmentally Friendly Agriculture, Chonnam National University) ;
  • Park, Ae Ran (Department of Agricultural Chemistry, Institute of Environmentally Friendly Agriculture, Chonnam National University) ;
  • Kim, Jin-Cheol (Department of Agricultural Chemistry, Institute of Environmentally Friendly Agriculture, Chonnam National University) ;
  • Kim, Soonok (Microorganism Resources Division, National Institute of Biological Resources) ;
  • Park, Sook-Young (Department of Plant Medicine, Sunchon National University)
  • 투고 : 2021.02.22
  • 심사 : 2021.07.21
  • 발행 : 2021.10.31
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초록

An endolichenic fungus Xylaria grammica EL000614 produces grammicin, a potent nematicidal pyrone derivative that can serve as a new control option for root-knot nematodes. We optimized an Agrobacterium tumefaciens-mediated transformation (ATMT) protocol for X. grammica to support genetic studies. Transformants were successfully generated after co-cultivation of homogenized young mycelia of X. grammica with A. tumefaciens strain AGL-1 carrying a binary vector that contains the bacterial hygromycin B phosphotransferase (hph) gene and the eGFP gene in T-DNA. The resulting transformants were mitotically stable, and PCR analysis showed the integratin of both genes in the genome of transformants. Expression of eGFP was confirmed via fluorescence microscopy. Southern analysis showed that 131 (78.9%) out of 166 transformants contained a single T-DNA insertion. Crucial factors for producing predominantly single T-DNA transformants include 48 h of co-cultivation, pretreatment of A. tumefaciens cells with acetosyringone before co-cultivation, and using freshly prepared mycelia. The established ATMT protocol offers an efficient tool for random insertional mutagenesis and gene transfer in studying the biology and ecology of X. grammica.

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과제정보

This work was supported by the National Institute of Biological Resources, funded by the Ministry of Environment of the Republic of Korea (projects NIBR201921101 and NIBR202021101) and by the Ministry of Science and ICT, Korea, under the Grand Information Technology Research Center support program [IITP-2021-2020-0-01489] supervised by the IITP. The USDA National Institute of Food & Agriculture and Federal Appropriations (Project PEN04655; Accession # 1016291) supported S. Kang

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