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Qualitative and Quantitative Assessment of Plasmid DNA Isolation, Rev A

oximately 8 x 108 cells/ml


Results
A wide variety of E. coli strains are frequently used for the propagation of plasmids. We have found that certain bacterial strains and certain plasmid purification kits can result in poor plasmid yield or poor plasmid quality. Therefore, we assessed the yield and quality of plasmids isolated from 14 common laboratory E. coli strains with the new Aurum miniprep and midiprep kits. We tested both endA- and endA+ bacterial strains because both the quality and quantity of plasmid DNA isolated from endA+ strains can be poor. The yields from these experiments are summarized in Tables 1 and 2. On average, ~20 g and 120 g of plasmid were isolated from minipreps and midipreps, respectively. The yields from minipreps varied more than from midipreps, but this may be because more variables (strains, endA- vs. endA+) were tested. In general, yield correlated most closely to specific plasmid type. Notably, good yields of plasmid could be obtained from endA+ strains.

Bacterial growth media can affect the quality and quantity of isolated plasmid DNA. At least four different media are commonly used for the growth of E. coli. We routinely use LB Amp but were interested in comparing the plasmid yield and quality when LBG Amp, 2x YT, or TB Amp was used. We found that plasmid yield seems to vary considerably with the type of medium used (Table 3). Notably, LBG Amp medium gave poor yields and plasmids that migrated unusually on agarose gels (Table 3, Figure 1). LB Amp and TB Amp gave the best yields and highest quality of plasmid.

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