One unit of Platinum® Taq Antibody is the amount of product required to inhibit one unit of Taq DNA polymerase from incorporating deoxyribonucleotide into acid-precipitable material.
20 mM Tris-HCl (pH 8.0), 40 mM NaCl, 2 mM Sodium Phosphate, 0.1 mM EDTA, 1 mM DTT, stabilizers, 50% (v/v) glycerol.
The Certificate of Analysis (CofA) provides detailed quality control information for each product. The CofA is available here, and is searchable by product lot number, which is printed on each box.
Since PCR is a powerful technique capable of amplifying trace amounts of DNA, all appropriate precautions should be taken to avoid cross-contamination. Ideally, amplification reactions should be assembled in a DNA-free environment. Use of aerosol-resistant barrier tips is recommended. Take care to avoid contamination with the primers or template DNA used in individual reactions. PCR products should be analyzed in an area separate from the reaction assembly area.
- Measure out the amount of Taq DNA polymerase needed. When using a reaction cocktail, determine the total number of units in the mix.
- Add an amount of “equivalent units” of Platinum® Taq Antibody to an equal amount of Taq DNA polymerase units. For Taq DNA polymerase at 5 unit per μl, prepare a 1:1 mixture. Mix well using a Vortex Mixer.
Add the following components to a sterile 0.5-ml microcentrifuge tube:
10X PCR Buffer, Minus Mg 5 μl 1X
10 mM dNTP mixture 1 μl 0.2 mM each
50 mM MgCl2
1.5 μl 1.5 mM
Primer mix (10 μM each) 1 μl 0.2 μM each
Platinum® Taq Antibody:
Taq DNA polymerase
2.5 units (or as required)
Autoclaved, distilled water
to 50 μl
If desired, a master mix can be prepared for multiple reactions to minimize reagent loss and to enable accurate pipetting.
- Mix contents of the tubes and overlay with 50 μl of mineral or silicone oil, if necessary.
- Cap the tubes and centrifuge briefly to collect the contents
- Incubate tubes in a thermal cycler at 94°C for 30 s to 2 min to completely denature the template and activate the enzyme.
- Perform 25-35 cycles of PCR amplification as follows: Denature 94°C for 30 s Anneal 55°C for 30 s Extend 72°C for 1 min per kb
- Maintain the reaction at 4°C after cycling. The samples can be stored at -20°C until use.
- Analyze the amplification products by agarose gel electrophoresis and visualize by ethidium bromide staining. Use appropriate molecular weight standards
- Chou, Q., et al. (1992) Nucl. Acids Res., 20, 1717.
- Sharkey, D.J., et al. (1994) BioTechnology, 12, 506.
- Westfall, B.A., et al. (1997) Focus®, 19.3, 46.
- Barnes, W.M. (1992) Gene, 112, 29.
- Lawyer, F.C., et al. (1993) PCR Methods and Applications, 2, 275.