Product Class: Other

Q5® Blood Direct 2X Master Mix
cloned at NEB recombinant No extraction-free PCR hifi hotstart pcr 3


Catalog #M0500

Product Introduction

  • Skip DNA purification and amplify direct from human whole blood cells or dried blood spots
  • Utilize Q5 Hot Start High-Fidelity DNA Polymerase for accurate and efficient PCR
  • Benefit from increased resistance to inhibitors and chemicals with our optimized buffer system
  • Set up reactions at room temperature with aptamer-based enzyme control

Product Information

Description

Q5 Blood Direct 2X Master Mix can amplify a wide variety of targets direct from dried blood spots or up to 30% whole human blood, skipping DNA purification. The master mix includes Q5 Hot Start High-Fidelity DNA Polymerase, a thermostable DNA polymerase, with 3′→ 5′ exonuclease activity, fused to a processivity-enhancing Sso7d domain and dNTPs in an optimized buffer with increased resistance to inhibitors in blood, anti-coagulants, and chemicals on filter papers. It is capable of amplifying products up to 7.5 kb from human whole blood cells preserved with sodium EDTA, potassium EDTA, sodium citrate and sodium heparin as well samples stored on common preservative filter papers.

Q5 Hot Start High-Fidelity DNA Polymerase offers robust DNA amplification with low error rates. The addition of an aptamer-based inhibitor allows for room temperature reaction setup.

It is recommended to use NEB’s Tm Calculator to determine the appropriate annealing temperature for each primer pair and polymerase/buffer of interest. Unlike other calculators, the NEB Tm Calculator takes buffer components that affect melting temperatures into consideration when calculating the best annealing temperature. Other online calculators may underestimate the best Q5 annealing temperature.

Figure 1: Q5 Blood Direct 2X Master Mix enables robust amplification direct from whole or dried blood

Figure 1

PCR was performed using Q5 Blood Direct 2X Master Mix under standard recommended conditions with 35 cycles of amplification. Yield and purity were quantitated by microfluidic LabChip® analysis and are indicated by dot size and color, respectively, with a large, dark green dot representing the strongest performance. A. Amplification of a variety of human genomic amplicons, 0.3 to 7.5 kb in length, from 10% EDTA-preserved human whole blood. Results are shown as both a virtual gel (top) and corresponding dot plot (bottom). Q5 Blood Direct 2X Master Mix performs well across a broad range of amplicon sizes. B. Amplification of a 604 bp human genomic amplicon from whole blood (top) or blood dried on filters (bottom). Human whole blood comprised 5-30% of the total reaction volume (50 µl) as indicated. Untreated 1 mm punches from dried blood spots were added directly to 25 µl reactions (one punch per reaction), even where pre-treatment of the punch was recommended by the manufacturer. Q5 Blood Direct 2X Master Mix shows broad tolerance to varying blood volumes, preservatives and punch types.




Figure 2: Q5 Blood Direct 2X Master Mix offers strong performance on the widest variety of targets

Figure 2

PCR was performed with human amplicons varying in (A) GC content (23-77%) and (B) length (2 to 7 kb), using Q5 Blood Direct 2X Master Mix and several other commercially available blood direct polymerases. 10% EDTA-preserved human whole blood was used as starting material input. 50 µl reactions were cycled according to manufacturers’ recommendations. Yield and purity were quantitated by microfluidic LabChip® analysis and are indicated by dot size and color, respectively. Q5 Blood Direct 2X Master Mix shows consistently strong performance for the broadest range of targets.




Figure 3: Consistent performance across filter types, blood preservatives and reaction volumes



PCR was performed with a 604 bp human genomic amplicon from dried blood using Q5 Blood Direct 2X Master Mix under standard recommended conditions. Whatman 903 or FTA Elute paper was spotted with blood preserved with various anticoagulants. Punches from these dried blood spots were added direct to 20, 25 or 50 µl reactions, without pre-treatment. Amplification was robust regardless of volume size, anticoagulant, or filter type. 




 
This product is related to the following categories:
Q5® High-Fidelity DNA Polymerases,
Master Mixes,
PCR, qPCR & Amplification Technologies,
This product can be used in the following applications:
Extraction-Free PCR,
qPCR & RT-qPCR,
Hot Start PCR,
High-Fidelity PCR,
DNA Amplification, PCR & qPCR

Properties & Usage

Heat Inactivation

No

Protocols, Manuals & Usage

Protocols

  1. Protocol for Routine PCR Directly from Whole Blood or Dried Blood Spots (DBS) using Q5® Blood Direct 2X Master Mix (NEB #M0500)

Tools & Resources

Web Tools

FAQs & Troubleshooting

FAQs

  1. What is the difference between Q5® Blood Direct 2X Master Mix and Hemo KlenTaq (NEB #M0332)?
  2. What types of samples/materials are compatible with the Q5® Blood Direct 2X Master Mix?
  3. What concentration of whole blood is recommended for use with Q5® Blood Direct 2X Master Mix?
  4. What blood anticoagulants can be used with Q5® Blood Direct 2X Master Mix?
  5. How do I pre-treat Dried Blood Spot Punches?
  6. What length of product can be made by Q5® Blood Direct 2X Master Mix?
  7. What ends will my PCR products have?
  8. What are Hot Start and WarmStart® polymerases and when would I use them?
  9. What are the properties of this polymerase (fidelity, product ends, max amplicon, modified base incorporation, etc.)?
  10. My results are not as expected. Where can I find troubleshooting help?
  11. When and how should I use the Q5® High GC Enhancer?

Quality, Safety & Legal

Quality Assurance Statement

Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.

Specifications

The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]

Certificate Of Analysis

The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]

Safety DataSheets

The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

Legal and Disclaimers

Products and content are covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB). The use of trademark symbols does not necessarily indicate that the name is trademarked in the country where it is being read; it indicates where the content was originally developed. The use of this product may require the buyer to obtain additional third-party intellectual property rights for certain applications. For more information, please email busdev@neb.com.

This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.

Licenses

This product is covered by one or more patents. 

This product is licensed for research and commercial use from Bio-Rad Laboratories, Inc., under U.S. Pat. Nos. 6,627,424, 7,541,170, 7,670,808, 7,666,645, and corresponding patents in other countries. No rights are granted for use of the product for Digital PCR or real-time PCR applications, with the exception of quantification in Next Generation Sequencing workflows

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