Product Class: Other

T7 RNA Polymerase (High Concentration)
NEBU 37

GMP-grade reagent also available. Learn more.


Catalog #M0460

Product Introduction

In vitro transcription using T7 phage promoter 

Product Information

Description

T7 RNA Polymerase (High Concentration) is offered at a 20-fold higher concentration than our standard T7 RNA Polymerase (NEB #M0251) and is ideal for experienced users interested in building and optimizing their own in vitro transcription reactions. The enzyme is accompanied by RNA Polymerase Reaction Buffer and magnesium chloride solution to further enable testing of different conditions such as nucleotide concentrations. 

For standard RNA synthesis and high yield reactions, we recommend our T7 RNA Polymerase (NEB #M0251) and HiScribe®; kits. The HiScribe kits are specifically designed to produce high quality, high yield RNA in a short time with minimal optimization. 

In vitro transcription involves multiple components in addition to T7 RNA Polymerase including ribonucleotides, inorganic pyrophosphatase and RNase Inhibitor, which can be purchased separately. Please refer to the Related Products section for details. More information regarding transcription optimization can be obtained from the following article.  

Bacteriophage T7 RNA Polymerase is a DNA-dependent RNA polymerase that is highly specific for the T7 phage promoters. The 99 kD enzyme catalyzes in vitro RNA synthesis from a cloned DNA sequence under the T7 promoters. RNA produced using the T7 RNA Polymerase is suitable for many applications in research and biotechnology. 

Product Source

Isolated from E.coli carrying a plasmid which contains the T7 RNA Polymerase gene. 
This product is related to the following categories:
RNA Synthesis In vitro Transcription (IVT)
This product can be used in the following applications:
Transcription-Mediated and NASBA Amplification

Properties & Usage

Unit Definition

One unit is defined as the amount of enzyme that will incorporate 1 nmol ATP into acid-insoluble material in a total reaction volume of 50 µl in 1 hour at 37°C.

Reaction Conditions

1X RNAPol Reaction Buffer
Incubate at 37°C

1X RNAPol Reaction Buffer
40 mM Tris-HCl
6 mM MgCl2
1 mM DTT
2 mM spermidine
(pH 7.9 @ 25°C)

Storage Buffer

50 mM Tris-HCl
100 mM NaCl
20 mM β-ME
1 mM EDTA
50% Glycerol
0.1% (w/v) Triton® X-100
pH 7.9 @ 25°C

Unit Assay Conditions

1X RNAPol Reaction Buffer, supplemented with 0.5 mM each ATP, UTP, GTP, CTP, and 1 µg T7 DNA in 50 µl.

Application Features

  • mRNA for in vitro translation and micro injection 
  • Radiolabeled RNA probes
  • Non-isotopic RNA labeling
  • Preparation of RNA vaccines
  • Guide RNA for gene targeting
  • RNA structure, processing and catalysis studies
  • Anti-sense RNA for gene expression experiment 
  • RNA amplification
  • Isothermal amplification

References

  1. Yin, Y and Carter, C.W. (1996). Nucl. Acids Res. 24, 7, 1279–1286.
  2. Schenborn, E.T. and Meirendorf, R.C. (1985). Nucl. Acids Res. 13, 6223-6236.
  3. Davanloo, P., et al. (1984). Proc. Natl. Acad. Sci. USA. 81, 2035-2039.
  4. Sambrook, J., Fritsch, E.F. and Maniatis, T. (1989). Molecular Cloning: A Laboratory Manual. 2nd Ed, 10.27-10.37.
  5. Sambrook, J., Fritsch, E.F. and Maniatis, T. (1989).  Molecular Cloning: A Laboratory Manual. 2nd Ed, 18.82-18.84.
  6. Melton, D.A., et al. (1984). Nucl. Acids Res. 12, 7035-7056.
  7. Milligan, J.F., et al. (1987). Nucl. Acids Res. 15, 8783.
  8. Noren, C.J. et al. (1990). Nucl. Acids Res. 18, 83-88.
  9. Kreig, P.A. and Melton, D.A. (1984). Nucl. Acids Res. 12, 7057-7070.

Protocols, Manuals & Usage

Protocols

  1. Protocol for Standard RNA Synthesis

Application Notes

FAQs & Troubleshooting

FAQs

  1. What is the promoter sequence of T7 RNA Polymerase?
  2. Is it possible to start transcription with an A?
  3. Does the transcription reaction with T7 RNA Polymerase require a primer?
  4. Does T7 RNA Polymerase leave an extra base at the end of a transcript?
  5. Will T7 RNA Polymerase work on single stranded substrate?
  6. Will T7 RNA Polymerase work on uncut plasmid DNA?
  7. Can aberrant RNA be produced when using T7 RNA Polymerase?
  8. How can the yield of RNA be maximized when using T7 RNA Polymerase?
  9. Can I use T7 RNA Polymerase to make high specific activity radiolabeled probes?
  10. What are the main causes of reaction failure using T7 RNA Polymerase?
  11. Why is the specific activity of the probe low?

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 [email protected].

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.

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