Having carried out the PCR, products can be purified either by gel electrophoresis or using commercially available columns that remove primers, dNTPs and the PCR buffer. The products may then be cloned in one of the following ways:
T/A cloning. If a non-proofreading enzyme such as Taq polymerase is used then an additional adenine nucleotide is often added to the product at the 3' end. A linearised vector, T/A vector, has been developed with a 3' thymine overhang and this may be ligated to the PCR product;
Restriction Enzyme Addition: The PCR primers can have restriction enzyme sites included immediately before the sequence to be amplified. There must be some bases before the restriction site to enable the enzyme to recognise and cut the DNA. The PCR product and vector are then digested with the same enzyme and ligated;
Blunt ended ligation: Products made using enzymes such as Pfu that have proof-reading capabilities, and therefore have blunt ends, can be ligated directly into blunt-ended vectors.