Platinum GenoType Tsp DNA Polymerase

The Invitrogen Platinum GenoType Tsp DNA Polymerase is a recombinant DNA polymerase from a thermophilic bacterial species. Tsp DNA polymerase is a genetically engineered thermostable polymerase (Tsp) for genotyping of dinucleotide repeat loci. Platinum Tsp DNA polymerase is precomplexed with antibodies, which inhibits polymerase activity at room temperature. At approximately 72°C, the antibodies are denatured, and the enzyme is activated, allowing automatic hot-start PCR. Note that the Tsp antibodies are different from those used with Platinum Taq DNA Polymerase.

• Improved accuracy of allele size determination—Exhibits minimal non-templated nucleotide addition (also known as extendase activity); lacks both 5′ and 3′ exonuclease activities
• Convenient workflow—Can substitute for Taq DNA polymerase in PCR and allows room-temperature setup
• Increased specificity—Precomplexed with anti-Tsp antibodies for use in hot-start PCR

Tsp DNA polymerase is widely used for genotyping loci with dinucleotide repeats. Determining the size of amplified dinucleotide repeat regions can be challenging since Taq DNA polymerase often adds a non-templated nucleotide, such as one or more adenosines, to PCR products. The fraction of PCR amplicons containing an extra nucleotide is heterogeneous and is dependent on the sequence of the reverse primer used.

Platinum GenoType Tsp DNA polymerase displays reduced activity in adding an extra non-templated nucleotide to PCR products (Figure 1). This enzyme property enables researchers to determine the correct size of the alleles when genotyping loci with dinucleotide repeats by PCR.

Reverse primers modified with a 5´-GTGTCTT tail (also PIG-tail primers) are often used with Taq DNA polymerase to enhance the addition of extra nucleotides and to facilitate allele calling [1]. With these modified reverse primers, extra-nucleotide addition was increased but allele calling for the loci tested was not affected for Platinum GenoType Tsp DNA Polymerase. Therefore, Platinum GenoType Tsp DNA Polymerase is effective with either standard or modified reverse primers for PCR genotyping of dinucleotide repeat microsatellites (Table 1).

The Tsp DNA polymerase is not suitable for amplifying DNA fragments larger than 500 bp. The mutations that eliminate the 5′ exonuclease activity and decrease the extendase activity also diminish the enzyme’s ability to amplify long templates. Table 2 compares the properties of Tsp DNA polymerase and Taq DNA polymerase.

 Product info sheet for Platinum GenoType Tsp DNA Polymerase

1. Brownstein MJ, Carpten JD, Smith JR (1996) Modulation of non-templated nucleotide addition by Taq DNA polymerase: primer modifications that facilitate genotyping. Biotechniques 20(6):1004-1006, 1008-1000.