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Selection of parental monokaryons from Korean Hypsizigus marmoreus by protoplast regeneration

원형질체 재생을 통한 느티만가닥버섯 단핵균주 선발

  • Oh, Youn-Lee (Department of Biotechnology College of Life Sciences and Biotechnology Korea University) ;
  • Kong, Won-Sik (Mushroom Science Division, National Institute of Horticultural and Herbal Science, RDA) ;
  • Jang, Kab-Yeul (Mushroom Science Division, National Institute of Horticultural and Herbal Science, RDA) ;
  • Shin, Pyung-Gyun (Mushroom Science Division, National Institute of Horticultural and Herbal Science, RDA) ;
  • Kim, Eun-Sun (Mushroom Science Division, National Institute of Horticultural and Herbal Science, RDA) ;
  • Oh, Min ji (Mushroom Science Division, National Institute of Horticultural and Herbal Science, RDA) ;
  • Choi, In-Geol (Department of Biotechnology College of Life Sciences and Biotechnology Korea University)
  • 오연이 (고려대학교 생명공학과 대학원 생명공학과&BK21플러스) ;
  • 공원식 (농촌진흥청 국립원예특작과학원 버섯과) ;
  • 장갑열 (농촌진흥청 국립원예특작과학원 버섯과) ;
  • 신평균 (농촌진흥청 국립원예특작과학원 버섯과) ;
  • 김은선 (농촌진흥청 국립원예특작과학원 버섯과) ;
  • 오민지 (농촌진흥청 국립원예특작과학원 버섯과) ;
  • 최인걸 (고려대학교 생명공학과 대학원 생명공학과&BK21플러스)
  • Received : 2015.09.08
  • Accepted : 2015.09.30
  • Published : 2015.09.30

Abstract

Hypsizigus marmoreus is commercially the most important edible mushroom in Japan. This mushroom is usually cultivated for a longer period (about 85~120 days) than other mushroom. In order to develop a new cultivar that has a shortened cultivation period, the genome analysis of this strain has been considered. This study aims to obtain parental monokaryotic strains reproducing 'Haemi' cultivar in Hypsizigus marmoreus for reference genome sequencing. The mycelia were cultured in MCM and MYG media for various incubation periods. Homogenized mycelia were treated with commercial cell wall degrading enzymes to maximize protoplasts production yield from Hypsizigus marmoreus. The greatest number of protoplasts was obtained from mycelia cultured in MCM media for 3 days using Novozyme enzyme. The isolated protoplasts were grown in regeneration agar media after two weeks. Regenerated colonies were picked and moved on separated dishes for microscopic observation. Neohaplonts regenerated from dikayotic strains were identified by the absence of clamp connections. We confirmed that one of monokaryotic strains is a parental strain by crossing with an original compatible strain of 'Haemi' cultivar. This parental strain will be used for reference genome sequence analysis.

Keywords

Hypsigus marmoreus;Parental monokaryons;Protoplast isolation and regeneration

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