ExAct¢â Taq DNA Polymerase, High Fidelity
Applications
• PCR amplification of long DNA fragments up to 10 kb
• Extreme productivity
• High specificity and sensitivity
• RT-PCR
• Generation of PCR products for TA cloning
Description
ExActTM Plus Taq DNA Polymerase is an improved form of ExActTMTaq DNA Polymerase and can amplify target DNA at a broad-range of annealing temperature. Its unique buffer system reduces primer-dimer formation and non-specific amplification. ExActTM Plus Taq DNA Polymerase possesses the greater yield, processivity and fidelity than normal Taq polymerase. The fidelity of ExActTM Plus Taq DNA Polymerase is higher approximately 4 times than that of Taq polymerase. In case of PCR amplification of target DNA with high G+C content or structural problem, such as repeat sequence, the application of 5X Q Plus Buffer(optional) improves the specificity and productivity of the reaction.
PCR Reaction Conditions
PCR Reaction Conditions
25 ul volume 50 ul volume
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Template
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X ul
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X ul
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Forward Primer (10 pmoles/¥ìl)
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1 ul
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2 ul
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Reverse Primer (10 pmoles/¥ìl)
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1 ul
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2 ul
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5x Reaction Mix [with dNTPs]
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5 ul
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10 ul
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5x Q Plus Buffer(optional)
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0 ~ 10 ul
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0 ~ 20 ul
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ExActTM Plus
Taq DNA Polymerase
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1 ~ 1.25 units
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1 ~ 1.25 units
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Distilled water
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up to 25 ul
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up to 50 ul
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¡Ø 5x Q Plus Buffer is an additive altering the binding behavior of primer and template and can help the amplification that do not work well under standard PCR condition. Especially, 5x Q Plus Buffer can be used for the amplification of problematic template, such as high G+C content and repeat sequence regions. 5x Q Plus Buffer uses as adding into PCR reaction mixture from 0.5x to 2x.
Human genomic DNA
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10 ng – 100 ng
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Bacterial genomic DNA
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5 ng – 50 ng
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Purified plasmid or phage DNA
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1 ng – 5 ng
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Perform 25-40 cycles of PCR amplification as follows
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Initial Denaturation
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95¡É
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2 minutes
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25-40 cycles
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95¡É
55 -68 ¡É
72¡É
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20 seconds
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40 seconds
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50 seconds/kb
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Final Extension
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72¡É
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5 minutes
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Components
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250 units
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500 units
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1,000 units
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ExActTM Plus Taq DNA Polymerase
(1.25 units/ul)
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200 ul
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200 ul x 2 ea
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200 ul x 4 ea
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5x Reaction Mix
[with dNTPs]
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1 ml x 2 ea
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1 ml x 4 ea
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1 ml x 8 ea
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5x Q Plus Buffer
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0.5 ml
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0.5 ml x 2 ea
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0.5 ml x 4 ea
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Data Sheet
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1 ea
|
1 ea
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1 ea
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Product Specifications
Storage Buffer
50% Glycerol, 20 mM Tris-HCl (pH 8.0), 100 mM KCl, 0.1 mM EDTA, 1 mM DTT, 0.5% Tween 20, 0.5% Nonidet P-40, 1 mM PMSF
Storage and Stability
The ExActTM Plus Taq DNA Polymerase is shipped on Dry/Blue Ice. All kit components should be stored at -20¡É upon receipt. Excessive freeze/thawing is not recommended. When stored under optimum conditions, the reagents are stable for a minimum of 6 months from date of purchase.
Quality Control
ExActTM Plus Taq DNA Polymerase was passed from quality control assay for contamination of endo- or exodeoxyribonuclease and the bacterial host DNA host.
Safety Precautions
Harmful if swallowed. Irritating to eyes, respiratory system and skin. Please refer to the material safety data sheet for further information.
Unit Definition
1u is defined as amount of enzyme that required to catalyze the incorporation of 10 nmoles of dNTP into acid-insoluble material in 30 minutes at 72¡ÆC.
Note
• Recombinant ExActTM Plus Taq DNA Polymerase is the enzyme of choice for most PCR applications.
• The number of PCR cycles depends on the amount of template DNA in the reaction mix and on the expected yield of the PCR product. 25 -35 cycles are usually sufficient for the majority PCR reaction. Low amounts of starting template may require 40 cycles.