Purification of plasmid DNA by fast protein liquid chromatography on superose 6 preparative grade

We were able to reduce both the time and the use of hazardous chemicals associated with the previous plasmid isolation methods of high-pressure liquid chromatography and CsCl gradient centrifugation by employing fast protein liquid chromatography (FPLC). Plasmid was first crudely prepared from bacte...

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Veröffentlicht in:Analytical biochemistry 1989-03, Vol.177 (2), p.378-382
Hauptverfasser: McClung, J.Keith, Gonzales, Robert A.
Format: Artikel
Sprache:eng
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Zusammenfassung:We were able to reduce both the time and the use of hazardous chemicals associated with the previous plasmid isolation methods of high-pressure liquid chromatography and CsCl gradient centrifugation by employing fast protein liquid chromatography (FPLC). Plasmid was first crudely prepared from bacterial cultures by a standard alkaline lysis method. After an alcohol precipitation, the nucleic acids were divided into two equal portions. One half was used for a standard purification method employing CsCl centrifucation. The other was dissolved in FPLC buffer, treated with RNase A, and applied to a Superose 6 preparative grade column (HR 10 30 ). Plasmid eluted off the column within 20 min as a single, highly resolved peak. Plasmid isolated by FPLC had yields, purity, and transformation efficiencies similar to that isolated by CsCl centrifugation.
ISSN:0003-2697
1096-0309
DOI:10.1016/0003-2697(89)90069-9