With Mg2+ free buffer and separate Mg2+ solution, 250 Units
Optimase® Polymerase with Mg2+ free buffer and separate Mg2+ solution, 250 Units
With Mg2+ free buffer and separate Mg2+ solution, 250 Units
Optimase Polymerase is a thermostable DNA polymerase with an efficient 3′ to 5′ proof-reading exonuclease activity developed for unprecedented PCR performance in single nucleotide polymorphism (SNP) and mutation discovery studies. Initially developed for denaturing high performance liquid chromatography (DHPLC) research1-4, Optimase Polymerase is the enzyme of choice for other applications requiring high fidelity PCR amplification.
• SNP and mutation discovery and scoring with various techniques such as SURVEYOR® Nuclease genome scanning, DHPLC, SSCP, DGGE and others
• High fidelity cloning
- High fidelity proofreading polymerase. It has a 3’-5’ exonuclease activity.
- Enhanced Optimase Polymerase Buffer System
- Free online support and access to the automated Optimase ProtocolWriter™ for PCR protocol design and Optimase MasterMix Calculator™ found at www.MutationDiscovery.com™
- Offers superior PCR fidelity over other proofreading polymerases1-3.
- Enhances the life of the DNASep® Cartridge in DHPLC experiments; does not interfere with other downstream applications, and increases amplification robustness and fidelity
- Enables simple and automated generation of PCR protocols; thus minimizing PCR optimization.
- Muhr, D., Wagner, T. and Oefner, P. 2002. Polymerase chain reaction fidelity and denaturing high-performance liquid chromatography. J Chromatogr B Analyt Technol Biomed Life Sci, 782:105-110. Abstract Library Entry.
- Nickerson, M. 2003. “Influence of PCR on DHPLC Polymorphism Characterization.” In Hecker, K. (Editor): Genetic Variance Detection: The Nuts&Bolts of DHPLC in Genomics. 2003, DNA Press.
- Application Note #119u. 2002. ADS BIOTEC Optimase Polymerase delivers highest fidelity in PCR for WAVE® System analysis.
- Jagmohan-Changur, S., et al. 2003. EXO1 variants occur commonly in normal population: evidence against a role in hereditary nonpolyposis colorectal cancer. Cancer Res, 63:154-158. Abstract Library Entry.
- Mo, J.Y., Maki, H., and Sekiguchi, M. 1991. Mutation specificity of the dnaE173 mutator associated with a defect in the catalytic subunit of DNA polymerase III of Escherichia coli. J. Mol. Biol. 222: 925-936.
General Polymerase Questions:
Q. What is Optimase® Polymerase?
A. Optimase® Polymerase is a novel proof reading polymerase isolated from a thermophilic Archaeal bacterium.
Q. What are the advantages of Optimase® Polymerase versus the other polymerases on the market?
A. Optimase® Polymerase provides 3’ to 5’ exonuclease activity unlike Taq polymerases. This proofreading function allows for much higher fidelity of amplification. Optimase® Polymerase is also fully compatible with the WAVE® System DNASep® cartridges, whereas other proofreading polymerases and their buffers contain additives that shorten the lifetime of the DNASep® cartridge. Optimase® has been shown to have higher proof reading fidelity than other PCR high fidelity polymerases.
Reaction and Amplification Questions:
Q. Can I use Optimase® Polymerase with all of my existing reaction and thermocycling conditions?
A. The kinetics of DNA amplification with a proof reading enzyme are different from a non-proof reader DNA polymerase. Therefore, the PCR conditions are different. Use ProtocolWriter™ at www.mutationdiscovery.com to calculate the correct thermocycler protocol.
Q. Do I have to use the formula provided to calculate the annealing temperatures of my primer sets?
A. The Quick Start Guide included with each kit gives a formula that has been used to develop the algorithm for ProtocolWriter. For your convenience use ProtocolWriter, which is easily accessible at www.mutationdiscovery.com when calculating the PCR conditions for your experiments. Further optimization can be performed if necessary. Annealing temperatures calculated using other formulas are typically lower than those obtained using the Quick Start Guide recommended formula, which can impact the fidelity of the reaction and result in lower specificity.
Q. Why are longer extensions times sometimes recommended for Optimase® Polymerase?
A. Proofreading polymerases require time to remove misincorporated bases and replace with the correct nucleotide. For this reason Optimase® Polymerase may require slightly longer extension times.
Q. Is it necessary to vortex the tube with the enzyme before performing the reactions as stated on the tube label?
A. Yes. It is recommended that the polymerase be vortexed for three seconds prior to addition to the reaction vials or master mix to ensure even distribution of the polymerase and storage buffer. Additionally after the addition of Optimase® Polymerase, each reaction should be vortexed and briefly centrifuged prior to thermocycling.
Q. The enzyme was at room temperature (arrived with thawed ice in the box). Is it a problem?
A. Optimase® Polymerase is very temperature stable, and most likely will still perform well. Our studies have shown that there is no problem. However, if you experience any difficulties, please contact Technical Support.
Q Can I do Hot Start with Optimase® Polymerase?
A. Optimase® Polymerase is not a heat-activated polymerase, so there is no advantage to using a “hot start”. However, the polymerase activity will not be inhibited by most typical hot start protocols.
Q Can I use a buffer from another enzyme?
A. This is not recommended, especially when using with the WAVE® System.
Q What additives for the buffer are compatible with Optimase® Polymerase?
A. DMSO and betaine are compatible with Optimase® Polymerase and the DNASep cartridges as long as the following guidelines are followed. DMSO up to 10% is compatible with the DNASep cartridge. Betaine is compatible with our cartridges provided recommendations in our cartridge warranty document are followed and with a maximum final concentration between 1.25 and 2.5M. Always use active clean and more frequent hot washes.
Q Can I add a Taq polymerase to Optimase®?
A. Many researchers have successfully added Taq from various vendors to their PCR reactions with Optimase® Polymerase. Please contact customer support for protocols or information.
Q Are Optimase® Polymerase PCR amplified products compatible with TA cloning?
A. Optimase® Polymerase is a proof reading polymerase and it does not add A to the end of PCR amplified products in the same way as Taq polymerases. Therefore, Optimase® Polymerase is more suitable for blunt-end cloning, rather than TA cloning.
Q Can ProtocolWriter™ be used with other polymerases?
A. The algorithm has only been designed for Optimase® Polymerase. ADS BIOTEC does not guarantee successful use with other polymerases. Researchers have successfully used ProtocolWriter with GoTaq™ DNA Polymerase from Promega* and Taq DNA Polymerase from Sigma Aldrich.
*GoTaq™ DNA Polymerase is a trademark of Promega.