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Received April 30, 2009
Accepted June 1, 2009
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A simple and cost-effective method to prepare template DNAs for cell-free protein synthesis
Department of Fine Chemical Engineering and Chemistry, Chungnam National University, Daejeon 305-764, Korea 1Department of Chemical System Engineering, Hongik University, Jochiwon, Chungnam 339-701, Korea
dmkim@cnu.ac.kr
Korean Journal of Chemical Engineering, January 2010, 27(1), 179-182(4), 10.1007/s11814-009-0294-4
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Abstract
The scale-up of cell-free protein synthesis reactions involves the preparation of large amounts of template DNA. While ion-exchange column chromatography methods have commonly been used to obtain purified plasmid DNA for cell-free protein synthesis reactions, these methods are costly and difficult to expand to a large scale. In this work, we report that the routine isopropyl alcohol (IPA) precipitation method can be used to prepare cell-free-expressible DNA when the co-precipitated proteins are removed. Compared to column-purification procedures, the IPA-precipitation offers obvious advantages with respect to the cost and scaling-up of template preparation, and we believe that our finding will contribute to making cell-free protein synthesis system more practical for the rapid production of preparative amounts of recombinant proteins.
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Nakano H, Kawarasaki Y, Yamane T, Adv. Biochem. Eng. Biotechnol., 90, 135 (2004)
Sawasaki T, Ogasawara T, Morishita R, Endo Y, Proc. Natl. Acad. Sci. USA, 99, 14652 (2002)
Byun JY, Kim DM, Biotechnol. Bioprocess Eng., 13, 84 (2008)
Oh IS, Kim TW, Ahn JH, Keum JW, Choi CY, Kim DM, Biotechnol. Bioprocess Eng., 12, 574 (2007)
Calhoun KA, Swartz JR, Biotechnol. Prog., 21(4), 1146 (2005)
Kim TW, Keum JW, Oh IS, Choi CY, Park CG, Kim DM, J. Biotechnol., 26, 554 (2006)
Freitas SS, Santos JAL, Prazeres DMF, Biotechnol. Prog., 22(4), 1179 (2006)
Kim DM, Kigawa T, Choi CY, Yokoyama S, Eur. J. Biochem., 239, 881 (1996)
Kim DM, Choi CY, Ahn JH, Kim TW, Kim NY, Oh IS, Park CG, Biotechnol. Bioprocess Eng., 11, 235 (2006)
Kim YE, Kim DM, Choi CY, Biotechnol. Bioprocess Eng., 11, 258 (2006)
Ahn JH, Hwang MY, Oh IS, Park KM, Hahn GH, Choi CY, Kim DM, Biotechnol. Bioprocess Eng., 11, 420 (2006)
Birnboim HC, Doly J, Nucleic Acids Res., 7, 1513 (1979)
Schagger H, Jagow G, Anal. Biochem., 166, 368 (1987)
Lehrach H, Diamond D, Wozney JM, Boedtker H, Biochemistry, 16, 4743 (1977)