The Korean Journal of Mycology (한국균학회지)

The Korean Society of Mycology (한국균학회)
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Agrobacterium-mediated transformation using gill tissue of Flammulina velutipes

Agrobacterium을 이용한 팽이 버섯 주름조직의 형질전환

  • Park, Soon-Young (Mushroom Research Division, National Institute of Horticultural & Herbal Science, RDA) ;
  • Van Peer, Arend F. (Mushroom Research Division, National Institute of Horticultural & Herbal Science, RDA) ;
  • Jang, Kab-Yeul (Mushroom Research Division, National Institute of Horticultural & Herbal Science, RDA) ;
  • Shin, Pyung-Gyun (Mushroom Research Division, National Institute of Horticultural & Herbal Science, RDA) ;
  • Park, Yun-Hung (Mushroom Research Division, National Institute of Horticultural & Herbal Science, RDA) ;
  • Yoo, Young-Bok (Mushroom Research Division, National Institute of Horticultural & Herbal Science, RDA) ;
  • Park, Ki-Moon (Department of Food Science and Biotechnology, Sungkyunkwan University) ;
  • Kong, Won-Sik (Mushroom Research Division, National Institute of Horticultural & Herbal Science, RDA)
  • 박순영 (농촌진흥청 국립원예특작과학원 버섯과) ;
  • ;
  • 장갑열 (농촌진흥청 국립원예특작과학원 버섯과) ;
  • 신평균 (농촌진흥청 국립원예특작과학원 버섯과) ;
  • 박윤정 (농촌진흥청 국립원예특작과학원 버섯과) ;
  • 유영복 (농촌진흥청 국립원예특작과학원 버섯과) ;
  • 박기문 (성균관대학교 생명공학부 식품생명공학) ;
  • 공원식 (농촌진흥청 국립원예특작과학원 버섯과)
  • Received : 2010.06.04
  • Accepted : 2010.06.15
  • Published : 2010.06.30
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Abstract

Agrobacterium-mediated transformation was conducted in order to generate DNA insertional mutants of Flammulina velutipes. Agrobacterium tumefaciens AGL-1 harboring pBGgHg was transformed into gill tissues of Flammulina velutipes strain KACC42777. The transformants resistant on hygromycine ($30\;{\mu}g/ml$) were confirmed by PCR. The targeted insertional sites were amplified by inverse PCR and sequenced. To find the phenotype variation of all generated transformants, bottle cultivation which followed by the standard cultivation protocol were conducted. Color variation was observed on the cultivated fruiting bodies. Furthermore, the transformant pool will be used as a good genetic resources for studying gene function.

팽이버섯의 다양한 유전자의 기능을 연구하기 위한 방법으로 팽이 자실체의 주름조직을 사용하여 A. tumefaciens을 매개로 한 형질전환을 수행하였다. Hygromycin 항생제 저항성 유전자를 포함하고 있는 AGL-1 pBGgHg를 팽이버섯의 자실체 조직에 도입시킴으로서 형질전환체들을 얻을 수 있었다. 형질전환체 선발은 첫 번째로 hygromycin 선발 배지에서 선발한 뒤 gpd-FH, hph-R primer를 이용하여 PCR로 확인 할 수 있었다. hygromycin은 30 ug/ml에서 선발 효율이 가장 높았다. 형질전환체 중 일부는 갈색 자실체를 갖는 wild type과는 달리 백색 자실체를 나타내었다. 형질전환체의 southern blot결과 외래유전자가 팽이버섯 내부에 하나 또는 그 이상의 다양한 copy 수로 삽입되어 있음을 알 수 있었다. 또한 삽입된 유전자의 주변 염기서열에 대한 정보는 Inverse PCR을 통해 알 수 있었다.

Keywords

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