HomeFAQsCan PCR products be phosphorylated in the PCR mixture?
FAQ: Can PCR products be phosphorylated in the PCR mixture?
No, because the ammonium sulfate found in most PCR buffers inhibits T4 Polynucleotide Kinase (NEB #M0201). Instead, gel purify your DNA or use a G25 spin column. In a 50 µl reaction of 1X Kinase Reaction Buffer use up to 200 pmol of 5´-hydroxyl DNA termini with 1 mM ATP. Heat to 70°C for 5 minutes, then chill on ice. Add 20 units T4 Polynucleotide Kinase (NEB #M0201) and incubate at 37°C for 30 minutes. Alternatively, treat your primers with T4 Polynucleotide Kinase (NEB #M0201) prior to PCR amplification.
You have been idle for more than 20 minutes, for your security you have been logged out. Please sign back in to continue your session.
Sign in to your NEB account
Any items placed in your cart as a guest will be lost upon sign in or leaving the site.