Large-scale purification of plasmid DNA by fast protein liquid chromatography using a Hi-Load Q Sepharose column.
Anal Biochem
; 203(1): 169-72, 1992 May 15.
Article
em En
| MEDLINE
| ID: mdl-1524212
ABSTRACT
The large-scale purification of plasmid DNA was achieved using fast protein liquid chromatography on a Hi-Load Q Sepharose column. This method allows for the purification of plasmids starting from crude plasmid DNA, prepared by a simple alkaline lysis procedure, to pure DNA in less than 5 h. In contrast to the previously described plasmid purification methods of CsCl gradient centrifugation or high-pressure liquid chromatography, this method does not require the use of any hazardous or expensive chemicals. More than 100 plasmids varying in size from 3 to 15 kb have been purified using this procedure. A Mono Q Sepharose column was initially used to purify plasmids smaller than 8.0 kb; however, a Hi-Load Q Sepharose column proved more effective with plasmids larger than 8 kb. The loading of plasmids larger than 8 kb on the Mono Q column resulted in a high back pressure and the plasmid DNA could not be eluted from the column. Thus, for routine purification we utilize the Hi-Load Q Sepharose column. Plasmids purified by this method had purity, yield, and transfection efficiency in mammalian cells similar to those of plasmids purified by CsCl density gradient centrifugation.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Plasmídeos
/
DNA Bacteriano
/
Cloretos
/
Cromatografia Líquida
Idioma:
En
Revista:
Anal Biochem
Ano de publicação:
1992
Tipo de documento:
Article
País de publicação:
EEUU
/
ESTADOS UNIDOS
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ESTADOS UNIDOS DA AMERICA
/
EUA
/
UNITED STATES
/
UNITED STATES OF AMERICA
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US
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USA