PCR involves replication of a DNA template by a thermostable DNA polymerase. The processivity, specificity, and fidelity of the polymerase enzyme used can influence the efficiency, reproducibility, and yield of the PCR reaction. High-fidelity PCR, utilizes a DNA polymerase with a low error rate and results in a high degree of accuracy in the replication of the DNA of interest. Fidelity is critical when accurate sequence amplification of the gene target is needed, for example, when direct sequencing or cloning for downstream protein expression. Unwarranted mutation could severely impact your studies. Our analysis has shown that KOD enzymes are an easy choice for fast, accurate and high-yielding PCR. EMD Millipore's molecular biologists work to develop and formulate polymerases offering the highest specificity, fidelity and yield during PCR amplification. In addition, optimized buffer compositions, convenient master mixes and cycling parameters provide additional ease of use and data reproducibility. KOD DNA Polymerase (formerly KOD HiFi DNA Polymerase) is a recombinant form of Thermococcus kodakaraensis KOD1 DNA polymerase (Nishioka 2001). KOD is a high fidelity thermostable DNA polymerase that amplifies target DNA up to 6 kbp with superior accuracy and yield for PCR applications (Takagi 1997). The enzyme's 3'→5' exonuclease-dependent proofreading activity results in a lower PCR mutation frequency than any other commercially available DNA polymerase. The elongation rate and processivity are 5 times and 10 to 15 times higher, respectively, than for Pfu DNA polymerase, resulting in highly accurate and robust yield, in a short reaction time. The enzyme generates blunt-ended PCR products suitable for cloning with the Novagen Perfectly Blunt® and LIC Vector Kits.
Source
Recombinant Thermococcus kodakaraensis KOD1 DNA polymerase expressed in E. coli
Concentration
2.5 U/µl
Purity
> 90% homogeneous by SDS-PAGE
5′ Exonuclease
Less than 2% per unit of enzyme when incubated 1 h at 74°C in a reaction with 5′-labeled λ/Sca I digest
Nicking activity
None detected
Amplification effiency
Functional PCR
Storage
–20°C
*Manufactured by Toyobo and distributed by EMD Biosciences, Inc. Not available from EMD Biosciences, Inc. in Japan.
Note: Purchase of this product includes an immunity from suit under patents specified in the product insert to use only the amount purchased for the purchaser's own internal research. No other patents rights (such as 5' Nuclease Process patent rights) are conveyed expressly, by implication, or by estoppel. 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.
Catalogue Number71085 Brand Family Novagen® Features and benefits
Higher fidelity than Pfu DNA polymerase—excellent for cloning
Greater yield—extension speed is 2X faster than Taq DNA polymerase and 5X faster than Pfu DNA polymerase
Higher processivity—sequential nucleotide polymerization is 10- to 15-fold greater than Pfu and Tli DNA polymerases
Amplifies plasmid and lambda DNA templates up to 6 kbp
Amplifies genomic DNA templates up to 2 kbp
No truncated amplification products
References
References
Nishioka, M., et al. 2001. J. Biotechnol.88, 141. Takagi, M., et al. 1997. Appl. Environ. Microbiol.63, 4504.
Product Information
Unit of Definition
One unit is defined as the amount of enzyme that will catalyze the incorporation of 10 nmol of dNTP into acid-insoluble form in 30 min at 75°C, in a reaction containing 20 mM Tris-HCl (pH 7.5 at 25°C), 8 mM MgCl₂, 7.5 mM DTT, 50 µg/ml BSA, 150 µM each of dATP, dCTP, dGTP, dTTP (a mix of unlabeled and [<Sup>3</Sup>H]dTTP) and 150 µg/ml activated calf thymus DNA.
•
250 U
KOD DNA Polymerase (2.5 U/µl)
•
1 ml
10X Buffer #1 for KOD DNA Polymerase (pH 8.0)
•
1 ml
10X Buffer #2 for KOD DNA Polymerase (pH 8.8)
•
1 ml
25 mM MgCl₂
•
1 ml
dNTP Mix (2 mM each)
Declaration
Manufactured by Toyobo and distributed by Novagen. Not available from Novagen in Japan. Purchase of this product is accompanied by a limited license to use it in the Polymerase Chain Reaction (PCR) process for research use in conjunction with a thermal cycler whose use in the automated performance of the PCR process is covered by the up-front license fee, either by payment to Applied Biosystems or as purchased, i.e., an authorized thermal cycler.
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