Lab Manual
biological reagents
Cell Free Systems
In Vivo Systems
misc methods

Phosphorylation with PNK

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Polynucleotide Kinase (PNK) is used to add a phosphate to the 5' end of a DNA fragment.  We phosphorylate oligos for cassette mutagenesis and we phosphorylate primers for toe printing experiments (basically a fancy primer extension reaction we use to map the positions of ribosomes on an mRNA).

The best approach is to follow the directions provided by the manufacturer.  They usually provide buffer also.  But if it is not compatible with the next reaction you are planning this buffer works well also.

10 X PNK 10 X AB
0.5 M Tris-Cl pH 8 200 mM Tris-Cl pH 8
0.1 M MgCl2 100 mM MgCl2
50 mM DTT 500 mM NaCl
0.5 mg/mL BSA 10 mM DTT