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A simplified protocol for the semi-large scale recovery of plasmids from <i>Escherichia coli</i> grown on agar plates
Section of Gene Expression Regulation, Frontier Science Research Center, Kagoshima University, Kagoshima, Japan
Department of Pediatric Dentistry, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima, Japan
Department of Pediatric Dentistry, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima, Japan
Division of Basic Molecular Science and Molecular Medicine, School of Medicine, Tokai University, Kanagawa, Japan
Department of Surgery II, National Defense Medical College, Saitama, Japan
Department of Organ Regeneration, Graduate School of Medicine, Shinshu University, Nagano, Japan
Animal Genome Research Unit, Division of Animal Science, National Institute of Agrobiological Sciences, Ibaraki, Japan
- 1 Section of Gene Expression Regulation, Frontier Science Research Center, Kagoshima University, Kagoshima, Japan
- 2 Department of Pediatric Dentistry, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima, Japan
- 3 Department of Pediatric Dentistry, Graduate School of Medical and Dental Sciences, Kagoshima University, Kagoshima, Japan
- 4 Division of Basic Molecular Science and Molecular Medicine, School of Medicine, Tokai University, Kanagawa, Japan
- 5 Department of Surgery II, National Defense Medical College, Saitama, Japan
- 6 Department of Organ Regeneration, Graduate School of Medicine, Shinshu University, Nagano, Japan
- 7 Animal Genome Research Unit, Division of Animal Science, National Institute of Agrobiological Sciences, Ibaraki, Japan
Journal of Biomedical Science and Engineering·Volume 05 (2012)·Pages 406–408·Published 18 July 2012·DOI10.4236/jbise.2012.57051
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Abstract
Semi-large scale liquid cultivation of transformed Escherichia coli ( E. coli ) in medium (100-200 ml) has been widely used for the acquisition of relatively large amounts of plasmid DNA (50-300 μg). However, this method requires an expensive high-speed centrifugation apparatus to precipitate E. coli before lysis, which is both laborious and time-consuming. Here, we demonstrate a method for agar plate-based cultivation of bacteria that does not employ a high-speed centrifugation apparatus. This procedure proves to be simple and reproducible, yielding an average of 82 μg of plasmid DNA per experiment. It may therefore be valuable for cloning/transfection experiments under limited financial backgrounds.
KeywordsAgar PlateDNA Purification<i>E. coli</i>Gene EngineeringPlasmid
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