PWOSUP-RO

Pwo SuperYield DNA Polymerase

suitable for PCR, Difficult Templates/Specialty Enzymes PCR, hotstart: no, dNTPs included: no

• Enzyme Commission number: 2.7.7.7 (BRENDA, IUBMB)
• UNSPSC Code: 41106300
• NACRES: NA.55

Properties

• biological source: (pyrococcus woesei)
• Quality Level: 100
• form: liquid
• usage: sufficient for ≤200 reactions (04340850001); sufficient for 200 reactions; sufficient for ≤40 reactions (04340868001); sufficient for 40 reactions
• specific activity: 5 U/μL
• mol wt: ~ 90 kDa
• feature: Difficult Templates/Specialty Enzymes PCR; High Fidelity PCR; dNTPs included: no; hotstart: no
• packaging: pkg of 100 U (04340868001); pkg of 500 U (04340850001 [2 x 250 U])
• manufacturer/tradename: Roche
• concentration: 50 U/mL
• parameter: 72 °C optimum reaction temp.
• technique(s): PCR: suitable
• color: colorless
• input: purified DNA
• pH: 8.0 (39 °F)
• solubility: water: miscible
• suitability: suitable for PCR
• NCBI accession no.: AAB67984
• UniProt accession no.: P61876
• application(s): life science and biopharma
• foreign activity: Endonuclease 30 units, none detected; Nicking activity 30 units, none detected
• storage temp.: −20°C

Description

General description

Pwo SuperYield DNA Polymerase combines the recombinant enzyme Pwo DNA Polymerase with an optimized buffer system. This buffer system enhances the enzymatic properties of the polymerase, resulting in higher yields of the amplification reaction without changing the fidelity of DNA synthesis. This enzyme delivers excellent results due to its enzyme design and optimized buffer system. Amplify fragments up to 3 kb - even longer amplicons are possible from simple templates.
Pwo SuperYield DNA Polymerase exhibits increased thermal stability with a half life of greater than 2 hours at +100 °C compared to Taq DNA Polymerase with a half life of less than 5 min at this temperature.

Pwo SuperYield DNA Polymerase, originally isolated from Pyrococcus woesei, is a processive 5′ →3′ DNA polymerase with 3′ →5′ exonuclease proofreading activity; 5′ →3′ exonuclease activity is not detectable.
The enzyme accepts modified nucleotides for efficient labeling of nucleic acids by PCR.
PCR products are blunt-ended directly useable for blunt-end ligation.
Using the magnesium-containing reaction buffer supplied, the final MgCl₂ concentration is 1.5mM.

Specificity

Star activity: In cases where star activity is observed and/or the activity of the enzyme in the PCR mix is low, first purify the amplification product prior to the restriction enzyme digest using High Pure PCR Product Purification Kit.

Application

• Pwo Super Yield DNA Polymerase combines the recombinant enzyme Pwo DNA Polymerase with a newly optimized buffer system. Pwo SuperYield DNA Polymerase is used for the amplification of DNA with the intent to sequence the amplification product or to clone the product (e.g., for the expression of the gene product). The high fidelity of this enzyme makes it particularly suitable for: High fidelity PCR
• Site-directed mutagenesis
• Cloning
• Gene expression
• Study of allelic polymorphism in individual RNA transcripts
• Characterization of rare mutations in tissue
• Characterization of the allelic stage of single cells or single DNA molecules

Features and Benefits

Pwo SuperYield DNA Polymerase yields more high fidelity PCR product. The optimized buffer delivers superior results. Amplify fragments up to 3kb. A GC-RICH Solution is included for difficult templates.

• Higher yield and 18-fold higher fidelity

compared to Taq DNA Polymerase.

• High performance with difficult templates.

The GC-RICH Solution is for GC-rich PCR.

• Reduce working steps in cloning.

Perform digests directly in Pwo SuperYield PCR mix.

Packaging

1 kit containing 3 components

Quality

Each lot is assayed using activated DNA. PCR testing used λ and human genomic DNA.

Unit Definition

One unit Pwo SuperYield DNA Polymerase is defined as the amount of enzyme that catalyzes the incorporation of 10 nmol total deoxynucleoside triphosphates into acid precipitable DNA within 30 minutes at +70 °C.

Unit Assay: Incubation buffer for assay on activated DNA
20 mM Tris-HCl, pH 8.8 (20 °C), 50 mM KCl, 2.5 mM MgCl₂, 10 mM 2-mercaptoethanol, 0.2 mM of each dATP, dCTP, dGTP, dTTP.

Incubation procedure
12.5 mg activated calf thymus DNA and 0.1 mCi [α-³²P]dCTP are incubated with 0.01 to 0.1 U Pwo SuperYield DNA Polymerase in 50 μl incubation buffer with a paraffin-oil overlay at +70 °C for 30 minutes. The amount of incorporated dNTPs is determined by trichloroacetic acid precipitation followed by scintillation counting.

Volume Activity: 5 U/μl

Other Notes

Modified nucleotides are substrates
Pwo DNA Polymerase accepts modified nucleotides like digoxigenin-dUTP, biotin-dUTP, or fluorescein-dUTP. Thus, it can add these nucleotides to DNA during PCR. These nonradioactively labeled products can be used as a hybridization probe in many applications.

For life science research only. Not for use in diagnostic procedures.

Legal Information

Use of this product is covered by US patent claims and corresponding patent claims outside the US. The purchase of this product includes a limited, non-transferable immunity from suit under the foregoing patent claims for using only this amount of product for the purchaser′s own internal research. No right under any other patent claim, no right to perform any patented method, and no right to perform commercial services of any kind, including without limitation reporting the results of purchaser′s activities for a fee or other commercial consideration, is conveyed expressly, by implication, or by estoppel. This product is for research use only. Diagnostic uses under Roche patent claims require a separate license from Roche. Further information on purchasing licenses may be obtained by contacting the Director of Licensing, Applied Biosystems, 850 Lincoln Centre Drive, Foster City, California 94404, USA.</ br>NOTICE TO PURCHASER: LIMITED LICENSE<br />Use of this product is covered by US Patent No. 6,127,155 and corresponding patent claims outside the US.  The purchase of this product includes a limited, non-transferable immunity from suit under the foregoing patent claims for using only this amount of product for the purchaser′s own internal research.  No right under any other patent claim and no right to perform commercial services of any kind, including without limitation reporting the results of purchaser′s activities for a fee or other commercial consideration, is conveyed expressly, by implication, or by estoppel.  This product is for research use only. Human and veterinary diagnostic uses under Roche patent claims require a separate license from Roche.  All uses other than internal research and human and veterinary diagnostic uses under Roche patent claims require a separate license from Thermo Fisher Scientific. Further information on purchasing licenses from Roche may be obtained by contacting the Licensing Department of Roche Molecular Systems, Inc., 4300 Hacienda Drive, Pleasanton, California 94588, USA or Roche Diagnostics GmbH, Sandhofer Strasse 116, 68305 Mannheim, Germany.  Further information on purchasing licenses from Thermo Fisher Scientific may be obtained by contacting the Licensing Department of  Thermo Fisher Scientific, 5791 Van Allen Way, Carlsbad, California 92008, USA.

Kit Components Only

• Pwo SuperYield DNA Polymerase 5 U/μl
• PCR buffer, containing 15 mM MgSO₄ 10x concentrated
• GC-RICH Solution 5x concentrated

Safety Information

• Hazard Statements: H412
• Precautionary Statements: P273 - P501
• Hazard Classifications: Aquatic Chronic 3
• Storage Class Code: 12 - Non Combustible Liquids
• WGK: WGK 2
• Flash Point(F): does not flash
• Flash Point(C): does not flash