Korean Journal of Breeding Science (한국육종학회지)

The Korean Society of Breeding Science (한국육종학회)
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Modified CTAB DNA Methods for efficient DNA extraction from Rice (Oryza sativa L.)

벼 분자육종을 위한 CTAB DNA 추출 시스템 개량

  • 이종희 (농촌진흥청 작물과학원 영남농업연구소) ;
  • 곽도연 (농촌진흥청 작물과학원 영남농업연구소) ;
  • 여운상 (농촌진흥청 작물과학원 영남농업연구소) ;
  • 김춘송 (농촌진흥청 작물과학원 영남농업연구소) ;
  • 전명기 (농촌진흥청 작물과학원 영남농업연구소) ;
  • 강종래 (농촌진흥청 작물과학원 영남농업연구소) ;
  • 박동수 (농촌진흥청 작물과학원 영남농업연구소) ;
  • 신문식 (농촌진흥청 작물과학원 호남농업연구소) ;
  • 오병근 (농촌진흥청 작물과학원 영남농업연구소) ;
  • 황흥구 (농촌진흥청 작물과학원 영남농업연구소)
  • Received : 2008.09.09
  • Published : 2008.09.10
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Abstract

Many important traits have been tagged allowing plant breeders to apply marker assisted selection (MAS) in rice. PCR itself is simple to set up, and requires little hands-on time. However, a crucial limiting step of MAS programs is the reliable and efficient extraction of DNA which can be performed on thousands of individuals. In this study, We describe a modification of the DNA extraction method, in which cetyltrimethylammonium bromide (CTAB) is used to extract DNA from leaf tissues for suitable MAS in rice. We followed the standard 2% CTAB extraction method in all the procedure. In addition we used the 1.2 ml 8-strip tube instead of 1.5 ml E-tubes to fit the 8-multichannel pipette and employ the 96 well plate to use the swing bucket centrifuge. Our modified CTAB DNA extraction method offers several advantages with respect to traditional and simple methods. 1) adult leaf samples collected in paddy field are applicable. 2) 96 leaf samples can be homogenized only one-time by using tungsten carbonate bead and 96well block. 3) semiautomatic loading method using 8-multichannel pipette from DNA extraction to electrophoresis of PCR products. 4) our system can extract about 400 leaf samples per day by only one technicion. Therefore, this method could be useful for marker assisted breeding in rice.

본 연구에서 분자육종을 위한 신속 DNA추출법을 개발하고자, 2% CTAB 추출법을 시스템적으로 개량한 Modified CTAB법에 대한 결과를 요약하면 아래와 같다. 1. DNA 추출용 샘플마쇄는 Voltex mixer, 3개 텅스텐 구슬 및 96 deep well block을 이용하였으며, 5분 이내에 96개 샘플을 분쇄할 수 있었다. 2. Modified CTAB DNA 추출법은 1.5 ml E-tube 대신에 1.2 ml 8연식 tube를 사용하였으며 DNA 추출, PCR작업 및 전기영동까지 일련의 작업이 8채널 멀티피펫을 이용할 수 있는 시스템적으로 개선하였다. 3. Modified CTAB법은 어린잎 뿐 만 아니라 이앙 후 30일째 채취한 잎에서도 DNA 추출이 가능하기 때문에 기존의 육종포장에 재식된 재료를 활용할 수 있는 장점이 있다.

Keywords

References

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