Fast and accurate method for quantitating E. coli host-cell DNA contamination in plasmid DNA preparations.
Plasmid DNA is being used successfully as a gene delivery vector in a variety of clinical applications. Similar to other pharmaceutical products for clinical use, the plasmid vectors must meet rigorous purity standards. One important contaminant is the DNA of the host cell used to produce the plasmids. We have developed a new method to accurately quantitate E. coli host-cell DNA in plasmid preparations. This method is based on kinetic PCR using the ABI PRISM 7700 with 23S rDNA as a target. This precise assay is significantly faster and has a lower limit of quantitation than the currently used Southern-based methods.
Smith
G J
GJ
Althea Technologies, San Diego, CA, USA.
Helf
M
M
Nesbet
C
C
Betita
H A
HA
Meek
J
J
Ferre
F
F
D015139:Blotting, Southern; D004247:DNA / Q000032:analysis* / Q000302:isolation & purification; D004269:DNA, Bacterial / Q000032:analysis*; D004275:DNA, Ribosomal / Q000032:analysis; D004926:Escherichia coli / Q000235:genetics*; D008722:Methods; D010957:Plasmids / Q000235:genetics*; D016133:Polymerase Chain Reaction; D012338:RNA, Ribosomal, 23S / Q000032:analysis
D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D012338:RNA, Ribosomal, 23S; D004247:DNA