Thermolabile Proteinase K Typical Reaction Protocol
New England Biolabs
Published: 2022-02-10 DOI: 10.17504/protocols.io.bg9wjz7e
Abstract
Thermolabile Proteinase K is an engineered, subtilisin-related serine protease that will hydrolyze a variety of peptide bonds and is frequently used to cleanup enzymatic reactions or cell lysates.
- Heat inactivated following incubation at 55°C for 10 minutes.
- Optimal activity and stability for up to 24 months.
- Active in a wide range of reaction buffers with optimal activity between 20 - 40°C and pH 7.0 - 9.5.
- No detectable endonuclease, exonuclease, DNase or RNase contaminating activities.
Steps
1.
Reactions may be scaled-up linearly to accommodate larger amounts of substrate and larger reaction volumes. Optimal buffering reagents, enzyme quantity, incubation temperatures and times may vary for particular substrates. Typical restriction enzyme cleanup conditions are as follows:
To a 50µL containing 1µg and 10units of a restriction enzyme:
2.
Add 1µL, mix gently.
3.
Incubate reaction at 37°C for 0h 15m 0s.
4. Note: Optimal reaction buffers, reaction temperatures as well as additional reaction properties of Thermolabile Proteinase K can be found in detail in the Note: Optimal reaction buffers, reaction temperatures as well as additional reaction properties of Thermolabile Proteinase K can be found in detail in the FAQ. Optimization may be required for other cleanup applications (ligation, extension, PCR, etc.). Optimization may be required for other cleanup applications (ligation, extension, PCR, etc.)
Incubate reaction at 55°C for 0h 10m 0s to inactivate Thermolabile Proteinase K .
Note
