Speedy Supreme NZYTaq 2× Colourless Master Mix is a premixed ready-to-use solution containing Speedy Supreme NZYTaq DNA polymerase (MB390), a recombinant modified form of Taq DNA polymerase that combines hot-start like PCR capacity and a fast polymerization reaction. The enzyme is inactive at room temperature, avoiding extension of non-specifically annealed primers or primer-dimers and thus providing higher specificity and sensitivity of DNA amplification. The functional activity of the enzyme is restored during a short 5- minute incubation step at 95°C. The enzyme only requires 5 seconds to successfully amplify a DNA fragment of 1 kb. The enzyme retains its speed when amplifying fragments up to around 2 kb. Successful amplification of higher DNA fragments up to 6 kb in size can be reached using a 10 sec/kb extension step. Faster PCR can be further achieved by increasing the primers melting temperature, which increases PCR annealing temperature, thus allowing combining the annealing and extension PCR steps during PCR cycling (see below). The master mix contains dNTPs, reaction buffer and additives at optimal concentrations for the efficient amplification of a wide range of DNA templates. MgCl₂ final concentration is 2.5 mM, allowing the implementation of a variety of PCR protocols. For highly sensitive downstream applications it is recommended to purify the amplified PCR product using NZYGelpure (MB011) before usage in subsequent protocols. Supreme NZYTaq II DNA polymerase lacks 3’→5’ exonuclease activity. Resulting PCR products have an A-overhang and are suitable for cloning with NZYtech’s NZY-A PCR cloning kit (MB053) or NZY-A Speedy PCR cloning kit (MB137).