Platinum GenoType Tsp DNA Polymerase

Determining the size of amplified dinucleotide repeat regions can be challenging since Taq DNA polymerase often adds a nontemplated nucleotide to PCR products (e.g., one or more adenosines). The fraction of PCR amplicons containing an extra nucleotide is heterogeneous and is dependent on sequence of the reverse primer used.

Platinum Genotype Tsp DNA polymerase displays reduced activity in adding an extra nontemplated nucleotide to PCR products. 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 addition of extra nucleotides and to facilitate allele calling [1]. With these modified reverse primers, extranucleotide 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.

The following table compares properties of Tsp DNA polymerase and Taq DNA polymerase.

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.

Platinum Tsp DNA polymerase is precomplexed with antibodies, which are denatured at about 72°C. Denaturation of the antibodies in the first step of PCR activates the enzyme and warrants automatic hot-start PCR. Note that the Tsp antibodies are different from those used with Platinum Taq 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.