Platinum Genotype Tsp DNA Polymerase
Invitrogen™ Platinum™ GenotypeTsp DNA Polymerase is a unique genetically engineered thermostable polymerase (Tsp) which is precomplexed with anti-Tsp antibodies for use in Hot Start PCR. It is designed specifically and functionally tested for amplification of dinucleotide repeat loci in genotyping reactions. It is genetically engineered such that:
- It does not have any 3′ exonuclease activity.
- It lacks 5′ exonuclease activity.
It's extendase activity (tendency to tail DNA with "nontemplate additions") is significantly diminished. Platinum Genotype Tsp DNA polymerase is engineered so that it can be readily substituted into standard PCR conditions that were designed for Taq DNA polymerase. PCR reactions employing Platinum Tsp can be set up at room temperature.
Properties of Tsp Polymerase vs. Taq DNA Polymerase
Researchers performing dinucleotide genotyping using Taq DNA polymerase often have difficulty determining the correct size of the alleles they are amplifying. This is due to the tendency of Taq to add non template nucleotides to PCR products. The PCR product fraction containing an extra nucleotide varies widely between primer sets and in any given reaction, the percentage of product containing an extra nucleotide may range from 0-100%. Platinum GenotypeTsp DNA polymerase addresses this problem because of its greatly reduced propensity for adding an extra nucleotide! In greater than 90% of the primer sets examined, PCR products exhibited no extra nucleotide addition, and where nontemplate nucleotide addition is detected, less than 10% of the PCR product actually contained the extra nucleotide.
|Features||Tsp DNA Polymerase||Taq DNA Polymerase|
|Molecular weight||70 kDa||94 kDa|
|Pigtail primers suggested for genotyping||No||Yes|
Limitations of Tsp
Tsp is NOT suitable for amplifying PCR fragments larger than 500 bp. (The same modifications which eliminated the 5′ exonuclease and changed the polymerase to decrease the extendase activity; decreases its ability to amplify large fragments).
Like Platinum Taq DNA Polymerase, Platinum Tsp DNA polymerase in precomplexed with antibodies. The antibodies, however, are different than those used with Platinum Taq. They denature at ~72°C, and are very effective for automatic Hot Start PCR.
|Detectable nontemplate Nt additions||Tsp DNA Polymerase||Taq DNA Polymerase|
|Primer sets tested||97||96|
|# with "n" pattern||86 (93%)||1 (1%)|
|# with "n/n+1" pattern||6 (7%)||34 (37%)|
|# with "n +1" pattern||0||56 (62%)|
|# with no amplification||5||5|