Recombinant Proteinase K Solution (20 mg/mL)

Applied Biosystems 7900HTの384-well blockで、FastモードのTaqMan Real-Time PCRを行う場合、User Bulletin(P/N 4369584) " Performing Fast Gene Quantitation with 384-Well Plates"に操作の詳細な指示がまとめられていますので、参照してください。

Scrape a single colony from the surface of an agar plate and transfer to 12 µL of SCL solution [10 mM Tris-HCl (pH 7.5), 1 mM EDTA, 50 µg/mL proteinase K] and incubate for 15 min at 55 degrees C. Inactivate proteinase K for 15 min at 80 degrees C and add 20 µL of deionized water. Centrifuge at 12,000 x g for 3 min. Transfer the supernatant to a new tube.

Concentration: Generally proteinase K is used in the concentration range of 50 to 500 µg/mL at 65 degrees C in the presence of SDS (0.5-1%).

Temperature optimum: 65 degrees C; 12X more active at 65 degrees C than at 25 degrees C.

pH: Proteinase K is stable over a wide pH range (4.0 to 12.5), with optimal activity at pH 6.5 to 9.5. It is most stable at pH 8.
Inactivation: Heat inactivate Proteinase K at 80 degrees C for 15 min. Phenol extraction is the recommended method to ensure complete inactivation.

Inhibited by: PMSF (0.1 to 1.0 mM of PMSF is usually sufficient for inhibition of Proteinase K)

Not inhibited by: Proteinase K is not inactivated by metal ions, chelating agents (e.g., EDTA), sulfhydryl reagents or by trypsin or chymotrypsin inhibitors. Activity can be stimulated by addition of denaturing agents (SDS and urea).

No, we do not recommend using it.

We do not sell the Proteinase K from the MagMAX mirVana Total RNA Isolation Kit as a standalone item. A different Proteinase K product (Cat. No. AM2548) can be purchased as a standalone item and can be used at 2.5X in the MagMAX mirVana protocol if you would like to add more Proteinase K to the procedure.