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Aurum Plasmid 96 Kit Tech Note, Rev A

m both 4 and 10 OD cultures were most similar to those of Aurum plasmid 96. However, Montage average concentrations were only about 60% of Aurum averages. Furthermore, the Montage protocol took about 80 min to complete, twice as long as the Aurum prep time. The protracted sample prep time for Montage was primarily due to an extremely slow plasmid binding step; wells containing lysates from 4 OD culture samples took 16 min to pass through the plasmid binding plate, as opposed to the 7 min indicated in the manual. Washing the plasmid binding plate required extra time as well. Plasmid recovery consisted of a manual transfer of resuspended DNA into a 96-well microplate via micropipet, as required in the protocol. The extent of handling in this step was consequently greater than in vacuum elutions.

Concert(96) Kit
The Concert(96) kit offered the user the opportunity to perform the lysis without centrifuging the bacteria beforehand (the direct load method), saving time. Any time saved, however, was lost by the lengthy centrifugations required to filter the plasmid samples, extending the plasmid preparation time to 70 min. Although plasmid sequencing quality was good, the lysate filtration and washes were unsatisfactory. Visible precipitates were observed in approximately 30 of 96 resuspended plasmid samples. The aberrant fluorescence observed on the agarose gel (Figure 2A) was probably caused by such precipitates. The low average A260/A280 ratios obtained with Concert(96) (ratios of 1.44 and 1.23) gave further evidence of sample impurity.


Conclusions
Obtaining high-quality DNA sequencing results is typically the primary criterion for choosing
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