Multiplex PCR NZYTaq 2x Green Master Mix

Storage Conditions:

-85°C to -15°C

Multiplex PCR NZYTaq 2× Green Master Mix is a premixed ready-to-use solution designed for the simultaneous amplification of multiple DNA fragments (up to 15 targets) in a single tube.

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Multiplex PCR NZYTaq 2× Green Master Mix is a premixed ready-to-use solution designed for the simultaneous amplification of multiple DNA fragments (up to 15 targets) in a single tube. The Master Mix contains dNTPs and reaction buffer at optimal concentrations for efficient multiplexing of targets ranging in size from 70 bp to 2.5 kb over a broad range of primer and template concentrations. MgCl₂ final concentration is 2.5 mM, allowing the implementation of a comprehensive variety of PCR protocols. Amplification efficiency is further improved by the inclusion of a recombinant modified enzyme derived from Taq DNA polymerase, which was engineered to display a hot-start-like PCR capacity and high processivity (fast polymerisation). The enzyme is inactive at room temperature, avoiding extension of non-specifically annealed primers or primer-dimers, and thus ensuring highly specific and sensitive multiplex PCR amplification. In addition, reactions assembled with Multiplex PCR NZYTaq 2× Green Master Mix may be directly loaded onto agarose gels, which simplifies the workflow. There are two dyes (blue and yellow) in the mix that allow monitoring the progress of electrophoresis. Multiplex PCR NZYTaq 2× Green Master Mix is not suitable when direct fluorescent or absorbance readings are required without prior purification of the amplified DNA from PCR. We recommend using the master mix version without dyes – Multiplex PCR NZYTaq 2× Colourless Master Mix (MB336) – or purifying the PCR product using NZYGelpure (MB011) before performing any other protocol.

Extension time: 30 sec/kb at 72 °C

3'→5' activity: no
Product overhang: 3'-A

More Information
Shipping Conditions	Blue Ice
Storage Conditions	-85°C to -15°C
Application	Multiplex PCR, TA Cloning, Typing/Detection
Features	Hot-start, Multiplex
Product Length	0-3 kb
Format	Mastermix
Product Category	End-Point PCR
Concentration	0.2 U/μL
Hot Start	Yes

Components

Multiplex PCR NZYTaq 2× Green Master Mix (0.2 U/μL)

More Information
Shipping Conditions	Blue Ice
Storage Conditions	-85°C to -15°C
Application	Multiplex PCR, TA Cloning, Typing/Detection
Features	Hot-start, Multiplex
Product Length	0-3 kb
Format	Mastermix
Product Category	End-Point PCR
Concentration	0.2 U/μL
Hot Start	Yes

FAQs

Multiplex PCR NZYTaq 2x Green Master Mix

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Multiplex PCR NZYTaq 2x Green Master Mix

What to do when there is no product amplification or low yield?

This can be due to the following situations:
a) Inadequate annealing temperature/ primer concentration. 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 (3 ° to 5 °C lower than T_m). Optimise annealing temperature using temperature gradient PCR. Increase primer concentration for lower yield amplicons.
b) Inadequate cycling conditions. Increase extension time and/or number of cycles.
c) 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.
d) Inadequate amount of template DNA. Titrate template amount. We recommend starting with 20-50 ng of genomic DNA.
c) Concentration of magnesium is too low. Mg²⁺ is included in the Master Mix at a final concentration of 2.5 mM, which is sufficient for most targets. For some targets, higher Mg²⁺ concentration may be required. Titrate from 2.5 mM to 4 mM (final concentration) in 0.25 mM increments. (Note: MgCl₂ is not provided in separate tubes).

How can I reduce the number of non-specific bands?

Adjust annealing conditions and/or design another pool of primers, avoiding complementary sequences. Ensure that each primer pair originates a single product without primer-dimers in individual PCR reactions. If necessary, reduce primer concentration. Reduce the number of cycles.

What is the maximum size of amplicon using Multiplex PCR NZYTaq 2x Green Master Mix?

Multiplex PCR NZYTaq 2x Green Master Mix allows efficient multiplexing of targets ranging in size from 70 bp to 2.5 k.

Which type of DNA ends are generated in the PCR products using Multiplex PCR NZYTaq 2x Green Master Mix?

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).

I don’t distinguish separate bands using agarose gel electrophoresis. What should I do?

When separating samples through agarose gel electrophoresis, please take into account that both concentration and grade of agarose are important when working with closely-sized fragments. NZYTech offers a vast portfolio of agaroses with different specifications to cover a wide range of needs. Please visit www.nzytech.com/products-services/category/molecular-biology/products/agaroses/ to identify the agarose that most suits your experiment.

How should I store my Multiplex PCR NZYTaq 2x Green Master Mix?

The product should be stored at -20 ºC, in a constant temperature freezer.

More Information
Shipping Conditions	Blue Ice
Storage Conditions	-85°C to -15°C
Application	Multiplex PCR, TA Cloning, Typing/Detection
Features	Hot-start, Multiplex
Product Length	0-3 kb
Format	Mastermix
Product Category	End-Point PCR
Concentration	0.2 U/μL
Hot Start	Yes

More Information
Shipping Conditions	Blue Ice
Storage Conditions	-85°C to -15°C
Application	Multiplex PCR, TA Cloning, Typing/Detection
Features	Hot-start, Multiplex
Product Length	0-3 kb
Format	Mastermix
Product Category	End-Point PCR
Concentration	0.2 U/μL
Hot Start	Yes

MSDS

Material Safety Data Sheets

MSDS EN MSDS PT

CoA

Certificate of Analysis

Manuals

Instructions for Use - EN

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