Abstract
Publisher Summary The cloning of amplified DNA from polymerase chain reactions (PCR) has become an important task for many researchers because the popularity of the PCR has increased. Unfortunately, the difficulty often encountered in cloning these fragments stands in marked contrast to their ease of production. Double-stranded DNA products obtained by primed amplification with Thermus aquaticus (Taq) DNA polymerase (and other polymerases that lack proofreading function) contain a single 3' untemplated deoxyadenosine residue gratuitously added by the polymerase to its product. To clone the amplified products efficiently, either the extra deoxyadenosine residues must be removed, or the vector has to be modified to accommodate them. It would be desirable to use the latter method in concert with a simple vector preparation, which would save the additional time and materials consumed in modifying the PCR product. A general method is described for direct cloning of DNA fragments generated by PCR that is based on digesting the cloning vector, pDK101, with Xcm I restriction endonuclease.
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