SupremeNZYLongDNA_pol

Supreme NZYLong DNA polymerase (MB331)

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Description: Supreme NZYLong DNA polymerase is an engineered version of NZYLong DNA polymerase designed to amplify longer PCR products, generally of 25 kb and beyond, which displays a higher specificity provided by its hot-start-like PCR capacity. This feature is achieved by a novel hot-start technology, which inhibits both polymerase and 3’→5’ exonuclease activities and thus avoids extension of non-specifically annealed primers or primer-dimers, as well as degradation of primers and template DNA during PCR reaction setup. The functional activity of the enzyme is restored during a short 5-minute incubation step at 94 °C. The increased processivity of Supreme NZYLong DNA polymerase combined with the hot-start-like PCR capacity results in higher specificity, sensitivity and yield during the amplification of long nucleic acids. A wide range of long PCR products, typically of 25 kb and up to 40 kb, can be generated usimg lambda DNA or human genomic DNA as starting template. Supreme NZYLong DNA polymerase displays a higher fidelity than conventional Taq DNA polymerases. The provided 10x reaction buffer protects DNA from depurination and nicking during long thermal cycling. The enzyme generates a mixture of A-overhang-ended (predominantly) and blunt-ended PCR products, being suitable for cloning with NZYTech´s TA PCR cloning kits (MB053 or MB137).

Features:
– Amplifies long DNA fragments
– High specificity: Eliminates nonspecific amplifications and minimizes primer-dimers
– Higher fidelity than NZYTaq DNA polymerase

Applications:
– Long-Range PCR
– Generation of products for TA cloning

 

Specifications:

Product length:0-20 kb
Hot-start like capacity:yes
Extension time:60 sec/kb at 68 °C
Product overhang:mixed
Available as Mixes:Supreme NZYLong 2x Green Master Mix (MB333); Supreme NZYLong 2x Colourless Master Mix (MB334)
Storage conditions:Store at -20 ºC
Shipping conditions:Shipped at 4 ºC to dry ice

Components:
– Supreme NZYLong DNA polymerase (5 U/μL)
– Reaction buffer (10x)

MSDS EN
MSDS PT
Product Brochure EN

1. What to do when there is no product amplification or low yield?
This can be due to the following situations:
a) Inadequate annealing temperature. The reaction mix composition may affect the melting properties of primers and DNA. Adjust the annealing temperature to accommodate the primer with the lowest melting temperature (5 ° to 10 °C lower than Tm).
b) Presence of PCR inhibitors. Some DNA isolation procedures, particularly genomic DNA isolation, can result in the co-purification of PCR inhibitors. Reduce the volume of template DNA in reaction or dilute template DNA prior to adding to the reaction. Diluting samples even 1:10,000 has been shown to be effective in improving results, depending on initial DNA concentration.
c) Template DNA damaged or degraded. An intact, high-quality template is essential to achieve a reliable amplification of large fragments. Extreme care must be taken in the preparation and handling of DNA. Always use purified high-quality DNA as template.
d) Concentration of magnesium is too low . Mg2+ is included in the 10× Reaction Buffer at a final concentration of 2 mM, which is sufficient for most targets. For some targets, a higher Mg2+ concentration may be required. Titrate from 2 mM to 4 mM (final concentration) in 0. 5 mM increments.

2. How can I reduce the number of non-specific bands?
Adjust annealing conditions and/or design another set of primers, by increasing the length and avoiding complementary sequences. Check the quality and concentration of primer solutions. We recommend to prepare small-volume working aliquots from the stock solution. Avoid using primers subjected to multiple freezing-thawing cycles.

3. How should I store my Supreme NZYLong DNA polymerase?
The product should be stored at -20 ºC, in a constant temperature freezer.

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