벼 분자육종을 위한 CTAB DNA 추출 시스템 개량
Modified CTAB DNA Methods for efficient DNA extraction from Rice (Oryza sativa L.)
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초록
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.
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.
참고문헌 (13)
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