The Korean Journal of Pesticide Science (농약과학회지)

The Korean Society of Pesticide Science (한국농약과학회)
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Characteristics of ${\delta}$-Endotoxin Protein Produced from Bacillus thuringiensis subsp. kurstaki KB099 Isolate Showing High Bioactivity against Spodoptera litura

담배거세미나방(Spodoplera lilura)에 높은 살충활성을 나타내는 Bacillus thuringiensis subsp. kurstaki KB099 균주의 내독소 단백질 특성

  • Jung, Sun-Young (Agro-Materials Safety Evaluation Division, National Academy of Agricultural Science Department of Agro-Food Safety, Rural Development Adminstration) ;
  • Seo, Mi-Ja (Dept. Applied Biology, College of Agriculture and Life Science, Chungnam National University) ;
  • Youn, Young-Nam (Dept. Applied Biology, College of Agriculture and Life Science, Chungnam National University) ;
  • Yu, Yong-Man (Dept. Applied Biology, College of Agriculture and Life Science, Chungnam National University)
  • 정선영 (농촌진흥청 국립농업과학원 농자재평가과) ;
  • 서미자 (충남대학교 농업생명과학대학 응용생물학과) ;
  • 윤영남 (충남대학교 농업생명과학대학 응용생물학과) ;
  • 유용만 (충남대학교 농업생명과학대학 응용생물학과)
  • Received : 2010.10.09
  • Accepted : 2010.11.30
  • Published : 2010.12.31
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Abstract

The characteristics of parasporal inclusion body from Bacillus thuringiensis subsp. kurstaki KB099 isolate which is high bioactive to the tobacco cutworm, Spodoptera litura, were examined. Parasporal inclusion of B. thuringiensis subsp. kurstaki KB099 isolate showed only 1 band at 130 kDa compared with B. thuringiensis subsp. kurstaki HD-l isolate producing 2 protein bands at 130 kDa and 60 kDa from by SDS-PAGE analysis without any enzyme treatment. Also, we confirmed that gut extract of sensitive S. litura KB099 isolate had digested only 60 kDa ${\delta}$-endotoxin protein. When the digestive enzyme of sensitive insect responsible for parasporal inclusion from KB099 and HD-l isolate was treated to each of them, protein band 60 KDa of KB099 was maintained up to 12 hours but all bands of HD-l were disappeared within 6 hours. In KB099 isolate, 6 genes (Cry1Aa, Cry1Ab, Cry1Ac, Cry1C, Cry1D and Cry1I) were identified by PCR analysis. Also, $Cry^-$ mutant of KB099 isolate was investigated by phase- contrast microscope, SDS-PAGE and PCR.

주요 농업해충인 담배거세미나방에 대하여 높은 생물활성을 보이는 Bacillus thuringiensis subsp. kurstaki KB099 균주의 내독소단백질의 특성이 검토되었다. 이 균주의 내독소단백질은 효소 처리 없는 SDS-PAGE 결과 HD-l 균주에서는 일부 용해되어 130 kDa과 60 kDa의 두 개의 단백질밴드가 나타났으나 KB099균주에서는 용해되지 않고 130kDa의 밴드만이 관찰되었다. KB099균주의 내독소단백질에 감수성 해충인 담배거세미나방의 중장액으로 소화하였을 때에는 약 60 kDa 단백질밴드가 형성됨을 확인하였다. 또한 두 균주의 각각의 내독소단백질에 감수성해충의 소화액으로 반응시켰을 때 생물활성이 약했던 HD-l균주는 약6시간 만에 주요 밴드가 사라지는데 비해 활성이 강한 KB099균주는 12시간이상 까지도 활성밴드가 유지되다가 24시간 정도에서 밴드가 사라졌다. KB099균주가 생산하는 내독소단백질 유전자의 탐색을 위한 PCR실험에서 Cry1Aa, Cry1Ab, Cry1Ac, Cry1C, Cry1D 그리고 Cry1I 등 6개의 유전자가 존재함을 확인하였다.

Keywords

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