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Rapid quantitation and monitoring of plasmid DNA using an ultrasensitive DNA‐binding dye
Author(s) -
Noites I. S.,
O'Kennedy R. D.,
Levy M. S.,
Abidi N.,
KeshavarzMoore E.
Publication year - 1999
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/(sici)1097-0290(1999)66:3<195::aid-bit8>3.0.co;2-6
Subject(s) - plasmid , plasmid preparation , alkaline lysis , recombinant dna , dna , escherichia coli , lysis , microbiology and biotechnology , chemistry , chromatography , fermentation , biology , fluorescence , biochemistry , gene , dna vaccination , physics , quantum mechanics , pbr322
A sensitive fluorescence‐based method for monitoring plasmid DNA during production was investigated. This simple method of assaying for plasmid DNA allows rapid monitoring of plasmid yields from a recombinant Escherichia coli fed‐batch fermentation. The assay has several advantages over traditional methods of plasmid DNA measurement. The fluorescent dye is highly specific and can measure total plasmid DNA concentration in about 5 min. The assay is sensitive over a wide range of plasmid concentrations of between 15 and 280 ng/mL, even in the presence of impurities that occur within alkaline lysate preparations. The technique can also be applied to monitoring fermentation and downstream purification steps. © 1999 John Wiley & Sons, Inc. Biotechnol Bioeng 66: 195–201, 1999.