Platinum™ Taq DNA Polymerase

Protocol tips

Upstream tips	Protocol tips	Downstream tips
Amplicon size should be consistent for each target sequence and limited to approximately 65 - 100 bp.	Always thaw on ice

Upstream tips
Amplicon size should be consistent for each target sequence and limited to approximately 65 - 100 bp.
Protocol tips
Always thaw on ice

Publication protocol

PCR Amplification

The primer set for the myotonic dystrophy assay was obtained from GeneLink (Catalog No.: 40-2026-10, Hawthorne, NY USA) and yields a product size of 114 + 3 N bp, where N is the number of triple-repeats. NEBNext® High-Fidelity 2X PCR Master Mix was provided by New England BioLabs (Catalog No.: M0541, Ipswich, MA USA). Each 25 μl reaction volume contained 0.5 μM of each primer and 1X NEBNext® High-Fidelity PCR Master Mix. Purified DNA or whole blood amounts for each reaction were used as described in the text. For Figure 1, 3 mM MgCl2 was required for amplification of the DM1 alleles from 30% whole blood. The standard 1X mix has 2 mM MgCl2, and increasing this to 3 mM did not improve amplification for other blood concentrations tested in this study. All reactions used MB grade nuclease-free water (Sigma, Catalog No.: W4502, St. Louis, MO USA). A 2-step PCR protocol was carried out using the Streck Philisa® Thermal Cycler (Catalog No.: 250000, Omaha, NE USA) and Philisa PCR Tubes (Catalog No.: 250005): Hot start of 98 °C for 3 minutes, followed by 30 cycles of [98 °C for 6 seconds and 68 °C for 12 seconds]. As a procedural control the Gene Link™ Myotonic Dystrophy Genemer™ kit (Catalog No.: 40-2026-11, Hawthorne, NY USA) was used as per manufacturer’s instructions, except Platinum® Taq DNA Polymerase (Life Technologies, Catalog No.: 10966-026, Grand Island, NY USA) was added to the PCR mix prior to the hot-start.


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Manufacturer protocol

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