PNK Treatment of DNA Ends

This step is used to add a phosphate group to the 5' end of a DNA molecule. Most primers for example are ordered without this being added as it requires an extra synthesis step and hence greater cost. However subsequent ligation steps are more efficient if these phosphate groups are added.

• T4 Polynucleotide Kinase (available from NEB)is an enzyme that can perform this on blunt or overhanging DNA ends.

• They describe a protocol for use which is summarized in a modified form below.