What is the function of proteinase K?
What is the function of proteinase K?
Proteinase K is used for the destruction of proteins in cell lysates (tissue, cell culture cells) and for the release of nucleic acids, since it very effectively inactivates DNases and RNases.
How do you inactivate proteinase K?
And the answer is very simple. Heat is a widely used way of inactivating proteinase K. While the activity of proteinase K increases with temperature, and is optimized at about 65 ˚C, heating proteinase K to 95 ˚C for 10 minutes will inactivate it.
What does buffer al do in DNA extraction?
The AL buffer contains the chaotropic agent guanidinium chloride. It promotes the lysis of the cell membrane, denaturation of proteins, DNA and other macromolecules. It also inactivates nucleases and promotes nucleic acid binding to pure silica material.
How do you make proteinase K buffer?
To ensure maximum Proteinase K activity, use water that has been treated with 0.1% DEPC (DEPC, GoldBio Catalog # D-340 [CAS 1609-47-8, mw. = 162.14 g/mol]) to make the buffer. Add 1 ml of DEPC to 1 L of H2O, shake vigorously, and let incubate for 12 hours at 37°C to inactivate any RNases.
Does proteinase K expire?
QIAGEN Proteinase K is stable for up to 1 year after delivery when stored at room temperature. To prolong the shelf-life of Proteinase K, storage at 2–8°C is recommended.
Can you add too much proteinase K?
Proteinase K is a protein and it will constitute a trivial amount of the protein in the extract and will be removed in the Ph-CHCl3 extraction. The SDS might hang around if you use too much, especially if you ethanol precipitate (though I suspect SDS is quite ethanol-soluble).
What does K stand for in proteinase K?
The enzyme was discovered in 1974 in extracts of the fungus Engyodontium album (formerly Tritirachium album). Proteinase K is able to digest hair (keratin), hence, the name “Proteinase K”.
What is QIAGEN proteinase K?
QIAGEN Proteinase K is a subtilisin-type protease isolated from the saprophytic fungus Tritirachium album and is particularly suitable for short digestion times. It possesses a high specific activity which remains stable over a wide range of temperatures and pH values with substantially increased activity at higher temperature.
What is proteinase K used for in PCR?
This PCR grade Proteinase K is extremely effective on native proteins and can therefore be used to rapidly inactivate endogenous nucleases such as RNases and DNases. This property makes Proteinase K particularly suitable for the isolation of native RNA and DNA from tissues or cell lines.
What is the normal range of Proteinase K?
~2.5 units/mg protein (Approximately 2.5 U/mg (Chromozym assay); ≥30 U/mg hemoglobin assay. Refer to the Certificate of Analysis for specific values for the present lot.) storage temp. Proteinase K is a subtilisin-related serine protease that has no pronounced cleavage specificity. The enzyme is also available as a lyophilizate.
What are the recommended buffer conditions for protease digestion?
‡ Recommended buffer conditions for protease digestion (Buffers B1+ B2) in the QIAGEN Genomic-tip protocol for bacterial DNA isolation. QIAGEN Proteinase K is a subtilisin-type protease isolated from the saprophytic fungus Tritirachium album and is particularly suitable for short digestion times.
