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

t was compared to PCR product purified using the silica-gel membrane method (figure 5). A 1.5-kb PCR product was generated using amplification primers, each containing a different restriction site sequence. The PCR product was digested with both restriction enzymes, purified using each method, ligated to digested pBluescript SK(+) vector and transformed into XL2-Blue ultracompetent cells. In five separate experiments, the 1.5-kb PCR product purified using the StrataPrep PCR purification kit and cloned showed an average transformation efficiency of 9.7 x 106 cfu/g of vector DNA. The percent of clones that contained inserts averaged 90%. For the 1.5-kb PCR product purified and cloned using the silica-gel membrane method, the transformation efficiency was 5.5 x 106 cfu/g of vector DNA, and the percent of clones that contained inserts averaged 82%. Thus, purification using the StrataPrep kit resulted in nearly a 2-fold increase in cloning efficiency and a higher percent of clones with inserts.

Gel Isolation

Figure 6

Figure 6 shows the yield of DNA fragments that were recovered from a conventional 1% agarose gel using either the StrataPrep DNA gel extraction kit or the silica-gel-membrane method. The two methods appear to be equivalent for fragments ranging in size from 0.25 to 9 kb. A higher yield of the 23-kb fragment is obtained with the StrataPrep DNA gel extraction kit. Transformation efficiencies for ligated 1.5- and 0.6-kb PCR products that were isolated with the StrataPrep DNA gel extraction kit were 1.7 and 9.8 x 106 cfu/g, respectively. These values were equival
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