Proteinase K |
Catalog No.GP10161 |
La proteasa K es una serina proteasa de amplio espectro y nuestro producto se extrae de células de Pichia pastoris con un gen clonado que codifica la proteasa endolítica de Engyodontium album (Tritirachium album).
Products are for research use only. Not for human use. We do not sell to patients.
Cas No.: 39450-01-6
Sample solution is provided at 25 µL, 10mM.
Cas No. | 39450-01-6 | M.Wt | 29.3 kDa |
Specific activity | >30 U/mg | Stability | pH range:4.0 to 12.5, temperature range: 25°C to 65°C. |
Solubility | Soluble in 20 mM Tris-HCl,1 mM CaCl2,50% Glycerol,pH 7.4 | Storage | Store at -20°C |
General tips | GlpBio guarantees optimal performance of this product for 18 months after date of delivery under the appropriate temperature and condition. | ||
Shipping Condition | Evaluation sample solution : ship with blue ice |
Quality Control & SDS
- View current batch:
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Specific activity: >30 U/mg
- Datasheet
This protocol is used in the extraction of genomic DNA from mouse tail with Proteinase K.
1. Place an approximately 0.5cm mouse tail into a 1.5 ml microcentrifuge tube.
2. Add 500 μl of lysis buffer (For example: 50 mM KCl, 50 mM Tris-HCl (pH 8.0), 2.5 mM EDTA,0.45% NP-40, 0.45% Tween-20).
3. Add 2.5 μl Proteinase K (20 mg/ml) to the tubeand mixthe solution.
4. Incubate overnight at 55°C.
?? Optional step:Incubate for an additional 1 hour at 65°C.
5. Vortex the tube and spin down for 10 seconds at 13,000 rpm to collect cell debris.
6. Use 1 μl from the top part of the supernatant per 50 μl of PCR mix(for genotyping).
Proteinasa K es una serina proteasa de amplio espectro y nuestro producto se extrae de células de Pichia pastoris con un gen clonado que codifica la proteasa endolítica de Engyodontium album (Tritirachium album). Es una proteasa altamente reactiva que se utiliza frecuentemente para digerir diversas proteínas y enzimas (incluyendo endonucleasas, exonucleasas, DNAsa o RNAsa). Por lo tanto, generalmente se usa en preparaciones de ADN sin afectar la integridad del ADN aislado. Tiene un rendimiento superior bajo una amplia gama de condiciones: pH, tampón, detergentes (como SDS), quelantes (como EDTA) y temperatura. La proteinasa K hidroliza los enlaces peptídicos preferentemente adyacentes al grupo carboxilo de aminoácidos hidrófobos (alifáticos, aromáticos y otros).
Además del aislamiento del genoma, también puede desempeñar una función en la detección de la localización de enzimas o en la eliminación de enzimas del ADN para mejorar la eficiencia de clonación.
La concentración de trabajo adecuada de la proteinasa K siempre está en el rango de 0,05 a 1 mg/mL. La actividad de la enzima puede ser estimulada por SDS del 0,2 al 1% o por urea del 1 al 4 mol. Se activa con calcio (1-5 mM), aunque los iones de calcio no afectan la actividad enzimática, sino que contribuyen a la estabilidad térmica y protegen a la proteinasa contra la autólisis. La proteinasa K tiene dos sitios de unión para Ca2+, que se encuentran cerca del centro activo pero no están directamente involucrados en el mecanismo catalítico. Por lo tanto, el ion calcio tiene una función reguladora para el sitio de unión del sustrato de la proteinasa K. La enzima es inactivada por DIFP o PMSF. Sin embargo, no es inhibida por EDTA, ácido iodoacético, inhibidor específico para tripsina TLCK, inhibidor específico para quimotripsina TPCK y p-cloromercuribenzoato.
Recomendamos un pH óptimo de 7,5 a 8,0 y una temperatura óptima entre 50 y 55°C. La desnaturalización rápida ocurrirá a temperaturas superiores a los 65°C. Puede mantenerlo bajo los 95°C durante 10 minutos como inactivación térmica.
References:
[1]. Kraus, E; et.al. Proteinase K from the Mold Tritirachium album limber, Specificity and Mode of Action. Z. Physiol. Chem., 357:939; 1976.
[2]. Jany,KD, et al. Amino Acid Sequence of Proteinase K from the Mold, Tritirachium albumlimber. Proteinase K; a Subtilisin-related Enzyme with Disulfide Bonds. FEBS Letter, 199,139.1986.
Cas No. | 39450-01-6 | SDF | |
Formula | M.Wt | 29.3 kDa | |
Solubility | Storage | ||
General tips | Please select the appropriate solvent to prepare the stock solution according to the
solubility of the product in different solvents; once the solution is prepared, please store it in
separate packages to avoid product failure caused by repeated freezing and thawing.Storage method
and period of the stock solution: When stored at -80°C, please use it within 6 months; when stored
at -20°C, please use it within 1 month. To increase solubility, heat the tube to 37°C and then oscillate in an ultrasonic bath for some time. |
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Shipping Condition | Evaluation sample solution: shipped with blue ice. All other sizes available: with RT, or with Blue Ice upon request. |
Step 1: Enter information below (Recommended: An additional animal making an allowance for loss during the experiment)
Step 2: Enter the in vivo formulation (This is only the calculator, not formulation. Please contact us first if there is no in vivo formulation at the solubility Section.)
Calculation results:
Working concentration: mg/ml;
Method for preparing DMSO master liquid: mg drug pre-dissolved in μL DMSO ( Master liquid concentration mg/mL, Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug. )
Method for preparing in vivo formulation: Take μL DMSO master liquid, next addμL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O, mix and clarify.
Method for preparing in vivo formulation: Take μL DMSO master liquid, next add μL Corn oil, mix and clarify.
Note: 1. Please make sure the liquid is clear before adding the next solvent.
2. Be sure to add the solvent(s) in order. You must ensure that the solution obtained, in the previous addition, is a clear solution before proceeding to add the next solvent. Physical methods such as vortex, ultrasound or hot water bath can be used to aid dissolving.
3. All of the above co-solvents are available for purchase on the GlpBio website.
Average Rating: 5
(Based on Reviews and 20 reference(s) in Google Scholar.)GLPBIO products are for RESEARCH USE ONLY. Please make sure your review or question is research based.
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