Ordering Information
- : RR001C
- : 3000 units
- : 5 units/μL
TaKaRa Ex Taq™ Polymerase
Description:
TaKaRa Ex Taq™ Polymerase
Trade Name:
Chemicon (Millipore)
Qty/Pk:
3000 units
Product Overview:
Ex Taq Polymerase is an optimized enzyme for long and accurate PCR. The enzyme is supplied at a concentration of 5 units/μL with a 10X Ex Taq Reaction Buffer and 2.5 mM dNTP Mix.
Unit definition: One unit is the amount of the enzyme that will incorporate 10 nmol of dNTP into acid-insoluble products in 30 minutes at 74°C with activated salmon sperm DNA as the template-primer.
Reaction mixture for unit definition: 25mM TAPS pH9.3 at 25°C; 50mM KCl; 2 mM MgCl2; 1mM 2-mercaptoethanol; 200μM each dATP, dGTP, dTTP; 100μM alpha P32-dCTP; 0.25mg/ml activated salmon sperm DNA
Unit definition: One unit is the amount of the enzyme that will incorporate 10 nmol of dNTP into acid-insoluble products in 30 minutes at 74°C with activated salmon sperm DNA as the template-primer.
Reaction mixture for unit definition: 25mM TAPS pH9.3 at 25°C; 50mM KCl; 2 mM MgCl2; 1mM 2-mercaptoethanol; 200μM each dATP, dGTP, dTTP; 100μM alpha P32-dCTP; 0.25mg/ml activated salmon sperm DNA
Key Applications:
DNA Amplification
Application Notes:
DNA amplification by Polymerase Chain Reaction (PCR).
PCR condition (an example)
When amplifying 1 kbp DNA fragment
30 cycles of : 98°C 10sec, 55°C 30sec, 72°C 1min or
30 cycles of 98°C 10sec, 68°C 1min. Note: Denaturation conditions vary depending on the cycler and tube. It is recommended for 10-30sec at 94°C, or 1-10sec at 98°C.
General reaction mixture for PCR (total 100μL)
TaKaRa Ex Taq™ 0.5μL
10X Ex Taq™ Buffer 10μL
dNTP Mixture (2.5mM each) 8μL
Template <1μg
Primer 1 20-100pmol (final conc. 0.2-1.0μM)
Primer 2 20-100pmol (final conc. 0.2-1.0μM)
Sterilized distilled water up to 100μL
PCR products: As most PCR products amplified with TaKaRa ex Taq™ have one A added at the 3'-termini, the obtained PCR product can be directly used for cloning into a T-vector. Also, it is possible to clone the product in blunt-end vectors after blunting and phosphorylation of the end.
Cool Start Method*: Minimizes the amplification of non-specific bands in pCR and achieves more accurate amplification. This is a simpler method without the need for special enzymes nor additional reagents.
Protocol:
1. Keep all reagents on ice until use.
2. Prepare the reaction mixture on ice. The adding order of reagents does not influence results. The result will not be affected even when the micutre is left on ice 30 min before thermal cycling.
3. Set a thermal cycler ready to start with the designated program. There is no need to change PCR conditions especially for Cool Start.
4. Set the tubes in thermal cycler and start cycling immediately.
* Japan patent 2576741 for Cool Start Method is owned by Shimadzu Corporation.
PCR condition (an example)
When amplifying 1 kbp DNA fragment
30 cycles of : 98°C 10sec, 55°C 30sec, 72°C 1min or
30 cycles of 98°C 10sec, 68°C 1min. Note: Denaturation conditions vary depending on the cycler and tube. It is recommended for 10-30sec at 94°C, or 1-10sec at 98°C.
General reaction mixture for PCR (total 100μL)
TaKaRa Ex Taq™ 0.5μL
10X Ex Taq™ Buffer 10μL
dNTP Mixture (2.5mM each) 8μL
Template <1μg
Primer 1 20-100pmol (final conc. 0.2-1.0μM)
Primer 2 20-100pmol (final conc. 0.2-1.0μM)
Sterilized distilled water up to 100μL
PCR products: As most PCR products amplified with TaKaRa ex Taq™ have one A added at the 3'-termini, the obtained PCR product can be directly used for cloning into a T-vector. Also, it is possible to clone the product in blunt-end vectors after blunting and phosphorylation of the end.
Cool Start Method*: Minimizes the amplification of non-specific bands in pCR and achieves more accurate amplification. This is a simpler method without the need for special enzymes nor additional reagents.
Protocol:
1. Keep all reagents on ice until use.
2. Prepare the reaction mixture on ice. The adding order of reagents does not influence results. The result will not be affected even when the micutre is left on ice 30 min before thermal cycling.
3. Set a thermal cycler ready to start with the designated program. There is no need to change PCR conditions especially for Cool Start.
4. Set the tubes in thermal cycler and start cycling immediately.
* Japan patent 2576741 for Cool Start Method is owned by Shimadzu Corporation.
Usage Statement:
Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
Quality Assurance:
PCR test: Good performance of DNA amplification by PCR was confirmed by using lamda DNA as the template (amplified fragment: 20kb).
Good performance of DNA amplification of beta-globin gene by PCR was also confirmed by using human genomic DNA as the template, and human p53-primers (amplified fragment: 17.5kb).
Good performance of DNA amplification of beta-globin gene by PCR was also confirmed by using human genomic DNA as the template, and human p53-primers (amplified fragment: 17.5kb).
Brand Family:
Chemicon
Sales Restrictions:
This product is only available for sale in the United States.
Presentation:
Taq Storage Buffer:
20mM Tris-HCL (pH8.0), 100mM KCl, 0.1mM EDTA, 1mM DTT, 0.5% Tween20, 0.5% Nonidet P-40, 50% glycerol
10X Ex Taq™ Buffer
1mL/vial
Mg2+ concentration (10X): 20mM
dNTP Mixture, ready for use in PCR without dilution:
800μL/vial
Concentration of 2.5mM of each dNTP
pH 7-9, dissolved in water with sodium salts. Purity greater than 98% for each dNTP
20mM Tris-HCL (pH8.0), 100mM KCl, 0.1mM EDTA, 1mM DTT, 0.5% Tween20, 0.5% Nonidet P-40, 50% glycerol
10X Ex Taq™ Buffer
1mL/vial
Mg2+ concentration (10X): 20mM
dNTP Mixture, ready for use in PCR without dilution:
800μL/vial
Concentration of 2.5mM of each dNTP
pH 7-9, dissolved in water with sodium salts. Purity greater than 98% for each dNTP
Storage Conditions:
Store supplied 10X buffer and dNTP mixture at -20°C.
Packaging:
12x250 units
Reagent Type:
PCR Enzymes
Protein or Enzyme Type:
- PCR Enzymes
- Polymerases
Product Name:
TaKaRa Ex Taq™ Polymerase
Concentration:
5 units/μL
Purity:
Nicking activity, endonuclease and exonuclease activity were not detected after the incubation of 0.6μg of supercoiled pBR322 DNA, 0.6μg of lamda DNA or 0.6μg of lamda-HindIII diegest with 10 units of this enyme for 1hr at 74°C.