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Simple Purification of DNA from Plasmid Minipreps, PCR Amplifications,,,and Agarose Gels

figure 3

Sequence analysis of purified nucleic acids is a rigorous functional test of DNA purity. DNA purified with the StrataPrep kit was sequenced using fluorescent dye terminator and dye primer chemistries with an ABI PRISM 377 sequencer (figure 3). In this data set, only 3 sequence ambiguities were identified out of approximately 600 bases. In a separate sequencing experiment, plasmid DNA was sequenced using a LI-COR 4000L DNA sequencer and infrared-labeled sequencing primers. Sequence comparisons were made of plasmid DNA purified using either the StrataPrep kit or a silica-gel membrane method. Both sequences were aligned with a published data set. For the silica-gel membrane method, 12 ambiguous base calls were identified in 801 bases of sequence. Only 8 ambiguities were seen in 795 bases of sequence from plasmid that was purified using the StrataPrep plasmid miniprep kit (data available upon request).

Purification of PCR Products

Figure 4

The StrataPrep PCR purification kit is effective at removing primers and nonspecific amplification products smaller than 100 bp. Figure 4 shows that ethidium bromide-stained material smaller than 100 bp is eliminated from 0.6- and 1.5-kb PCR products using the StrataPrep kit (lanes 3 and 7, respectively), whereas the silica-gel membrane retained contaminants migrating between 72 and 118 bp (lanes 4 and 8, respectively).

figure 5

Cloning efficiency of the 1.5-kb PCR product purified using the StrataPrep ki
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