Non-radioactive Phosphorylation with T4 PNK or T4 PNK (3´ phosphatase minus)
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|DNA||up to 300 pmol of 5´ termini|
|T4 PNK Reaction Buffer (10X)
|ATP (10 mM)
|T4 PNK||1 µl (10 units)|
|Nuclease-free Water||up to 50 µl|
2. Incubate at 37°C for 30 minutes.
3. Heat inactivate by incubating at 65°C for 20 minutes.
- ATP is not supplied
- 1X T4 DNA Ligase Buffer contains 1 mM ATP and can be substituted in non-radioactive phosphorylations (T4 Polynucleotide Kinase exhibits 100% activity in this buffer).