( PCR-Script cloning kits ...)Enhanced PCR Cloning System

PCR-Script cloning kits feature new additions

Tim Sanchez Julie Robinson Mary Buchanan
Stratagene

The PCR-Script cloning kits are convenient for cloning PCR fragments generated with any PCR enzyme.^1,2 The method is highly efficient, easy, and superior to other available methods.³ Now, Stratagene improves the kits by including both the StrataPrep PCR purification kit, so preparing PCR products is simple, and Epicurian Coli XL10-Gold ultracompetent cells,* so the highest transformation efficiencies of ligated DNA are attained. Furthermore, these innovative additions are available at no extra cost.

The core of these PCR-Script cloning kits is the PCR-Script cloning method,¹ in which PCR products are incubated for a 1-hour ligation reaction with one of the Srf I-digested PCR-Script vectors, Srf I restriction enzyme, and T4 DNA ligase. The Srf I enzyme recognizes the rare oligonucleotide sequence 5-GCCCGGGC-3. By including the Srf I enzyme in the ligation reaction, the PCR-Script cloning method maintains high concentrations of digested vector DNA and allows the use of nonphosphorylated, unmodified PCR products. This method ensures a low background of recircularized vector and increased ligation efficiency for blunt-ended DNA fragments by the simultaneous, opposing reactivity of the Srf I enzyme and T4 DNA ligase on nonrecombinant vector DNA. Ligated DNA is then ready to be transformed into competent cells.

The PCR-Script kits, although ideal to use with PCR enzymes such as Pfu and pfuturbo DNA polymerases,^,** are versatile and can be adapted for use with any PCR enzyme. Pfu and PfuTurbo DNA polymerases are proofreading enzymes that
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