Cloning Polymerase Chain Reaction (PCR) Products: Making T Vectors

Abstract

During polymerase chain reaction (PCR), DNA polymerases such as Taq add a single, unpaired residue—preferentially an adenosyl residue—to each 3′ end of a double-stranded amplified product. The unpaired terminal (A) residues can pair with a linear T vector that carries an unpaired 3′-thymidyl residue at each end. T vectors can be prepared in the laboratory or they may be purchased ready-made from commercial suppliers. This protocol outlines two methods in common use to generate T vectors.