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A simplified protocol for the semi-large scale recovery of plasmids from Escherichia coli grown on agar plates

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DOI: 10.4236/jbise.2012.57051    3,872 Downloads   6,450 Views   Citations

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.

Conflicts of Interest

The authors declare no conflicts of interest.

Cite this paper

Sato, M. , Akasaka, E. , Saitoh, I. , Ohtsuka, M. , Nakamura, S. , Sakurai, T. and Watanabe, S. (2012) A simplified protocol for the semi-large scale recovery of plasmids from Escherichia coli grown on agar plates. Journal of Biomedical Science and Engineering, 5, 406-408. doi: 10.4236/jbise.2012.57051.

References

[1] DNA Purification in Promega Home Page (http://www.promega.com/resources/product-guides-and-selectors/protocols-and-applications-guide/dna-purification/)
[2] QIAGEN Plasmid Purification System in Qiagen Home Page (http://www.ebiotrade.com/buyf/productsf/qiagen/QIAGEN_plasmid_purification_system.htm)
[3] Sambrook, J., Fritsche, E. and Maniatis, T. (1989) (Ed.), Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Press.
[4] Sato, M., Ishikawa, A. and Kimura, M. (2002) Direct injection of foreign DNA into mouse testis as a possible in vivo gene transfer system via epididymal spermatozoa. Molecular Reproduction and Development, 61, 49-56.
[5] Niwa, H., Yamamura, K. and Miyazaki, J. (1991) Efficient selection for high-expression transformants with a novel eukaryotic vector. Gene, 108, 193-200.
[6] van Ooyen, A., van den Berg, J., Mantei, N. and Weissmann, C. (1979) Comparison of total sequence of a cloned rabbit beta-globin gene and its flanking regions with a homologous mouse sequence. Science, 206, 337-344.

  
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