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At Bioline we pride ourselves in supplying high-quality polymerases to suit your requirements. To aid your selection of the most suited enzyme for your specific applications, please see our polymerase selection guide.
All our polymerases are guaranteed for a period of 12 months from the date of purchase. These should be stored at
-20°C during this time for optimal retention of activity.
Please Note: We do not recommend the storage of our polymerases at -80°C as ice crystals could form on the active site, which may affect or destroy the activity of the enzyme.
PCR can be a challenging technique, with various parameters to optimize to achieve the best results. If you are having problems, these could be easily resolved by addressing a few issues. Please see our PCR troubleshooting guide for suggestions and help with your specific problems.
These terms refer to parameters to be considered when performing PCR, and are important features in selecting the correct enzyme for your needs. Understanding what they mean is therefore crucial
- Yield: The amount of DNA produced in a PCR reaction
- Fidelity: The accuracy of the enzyme at incorporating the correct dNTP to the elongating DNA strand.
- Processivity: The length of time a polymerase is associated with the template, and therefore the size of fragment which can be amplified.
- Specificity: A measure of the unwanted by-products generated in a reaction.
In nature, the ability of a DNA polymerase to correct misincorporations of nucleotides in the DNA strand being elongated is often crucial to the survival of the host organism. This ability is termed “proofreading activity”, and occurs in the 3’ to 5’ direction. This activity also leads to the polymerase removing unpaired nucleotides overhanging at the 3’end (A-overhangs), creating blunt ends.
Please see the following table for the appropriate error rates:
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BIOLASE,
, IMMOLASE, Mango Taq, MyTaq DNA Polymerase
, Taq DNA Polymerase
| 1.1 x 10-4 base substitutions/bp (Tindall and Kunkel, 1988) |
| 2.4 x 10-5 frameshift mutations/bp (Tindall and Kunkel, 1988) |
| 2.1 x 10-4 errors/bp (Keohavang and Thilly, 1989) |
| 7.2 x 10-5 errors/bp (Ling et al., 1991) |
| 8.9 x 10-5 errors/bp (Cariello et al., 1991) |
| 2.0 x 10-5 errors/bp (Lundberg et al., 1991) |
| 1.1 x 10-4 errors/bp (Barnes, 1992) |
ACCUZYME |
1.6 x 10-6 errors/base (Lundberg et al., 1991). |
| VELOCITY |
4.4 x 10-7 errors/base (Frey & suppman, 1995) |
One unit is defined as the amount of enzyme that incorporates 10nmoles of dNTPs into acid- insoluble form in 30 minutes at 72°C.
All Bioline polymerases are available in the convenient format of a 2x master mix, containing all the reagents and additives necessary to perform successful PCR, with the exception of template, primers and water.
This formulation not only provides a convenient and hassle-free PCR setup, but also significantly reduces the chances of human error, inaccuracies and contamination by reducing the pipetting steps required.
All Bioline PCR mixes come supplied at a 2x concentration, requiring the user simply to add template, primers and PCR grade water to a final concentration of 1x.
We recommend a final reaction volume of 50µl, this requiring the use of 25µl of the 2x master mix, and to make up to 50µl using template, primers and PCR grade water.
Whilst the exact composition of the mixes is proprietary information, all mixes are made up of Reaction Buffer, Magnesium, dNTPs, Polymerase and additives, designed to keep the polymerase stable in the mix, as well as provide the optimal conditions for PCR. |
