Product Class: Other

ShortCut® RNase III
NEBU recombinant 37 No

Product Introduction

ShortCut RNase III converts long double-stranded RNA into a heterogeneous mix of short (18–25 bp) interfering RNAs (siRNA).

  • Isolated from a recombinant source
  • Useful for gene silencing and target validation
  • Supplied with 10X Reaction Buffer
  • Not sure which RNase to choose? Learn more about the different RNases offered by NEB.
Catalog # Size Concentration
M0245S 200.0 units 2000 units/ml
M0245L 1000.0 units 2000 units/ml

Product Information

Description

ShortCut® RNase III, used with its manganese-containing reaction buffer, converts long double-stranded RNA into a heterogeneous mix of short (18–25 bp) interfering RNAs (siRNA) suitable for RNA interference in mammalian cells (1–3). 1.5 units (1 µl) of ShortCut RNase III is sufficient to convert 1 µg of dsRNA into siRNA suitable for RNA interference in mammalian cells.

ShortCut RNase III digestion of dsRNA: (A) Varying amounts of ShortCut RNase III were incubated with 2 µg of a 500 bp dsRNA for 20 minutes at 37°C in a 50 µl reaction. Digests were analyzed by 20% TBE polyacrylamide electrophoresis. Marker lane contains a mixture of 21 bp siRNA Marker and 100 bp DNA Ladder (NEB #N3231). (B) dsRNA fragments (1 kb and 175 bp) were digested with ShortCut RNase III. Digests were analyzed by 20% TBE polyacrylamide gel electrophoresis.
GFP Silencing in COS-7 Cells: COS-7 cells co-transfected in a 24 well plate with a plasmid expressing GFP in the absence (control) or the presence of 30 ng (4 nM) of GFP siRNA prepared using the ShortCut RNAi Kit. Cells were photographed 48 hours post-transfection.

Product Source

An E. coli strain containing a genetic fusion of the E. coli RNase III gene (rnc) and the gene coding for maltose binding protein (MBP).
This product is related to the following categories:
RNases,
Epitranscriptome Analysis,

Reagents Supplied

Reagents Supplied

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration
  • M0245S     -20    
      ShortCut® RNase III M0245SVIAL -20 1 x 0.1 ml 2,000 units/ml
      ShortCut Reaction Buffer B0245SVIAL -20 1 x 1 ml 10 X
      10X EDTA B0255AVIAL -20 1 x 1 ml 500 mM
      MnCl2 B0786AVIAL -20 1 x 1 ml 200 mM
      Glycogen RNase-Free B1564SVIAL -20 1 x 0.05 ml 10 mg/ml
  • M0245L     -20    
      ShortCut® RNase III M0245LVIAL -20 1 x 0.5 ml 2,000 units/ml
      ShortCut Reaction Buffer B0245SVIAL -20 1 x 1 ml 10 X
      10X EDTA B0255AVIAL -20 1 x 1 ml 500 mM
      MnCl2 B0786AVIAL -20 1 x 1 ml 200 mM
      Glycogen RNase-Free B1564SVIAL -20 1 x 0.05 ml 10 mg/ml

Properties & Usage

Unit Definition

One unit is the amount of enzyme required to digest 1 µg of dsRNA to siRNA in 20 minutes at 37°C in a total reaction volume of 50 µl.

Reaction Conditions

1X ShortCut Reaction Buffer
Incubate at 37°C

1X ShortCut Reaction Buffer
50 mM Tris-HCl
1 mM DTT
50 mM NaCl
(pH 7.5 @ 25°C)

Usage Concentration

1X

Storage Buffer

500 mM NaCl
10 mM Tris-HCl
0.5 mM EDTA
1 mM DTT
50% Glycerol
pH 8 @ 25°C

Heat Inactivation

No

Unit Assay Conditions

One unit is the amount of enzyme required to digest 1 μg of dsRNA to siRNA in 20 minutes at 37°C in a total reaction volume of 50 μl.

Features

  • Make effective siRNAs against any gene target
  • Heterogeneous population of siRNA ensures silencing of target gene
  • From DNA template to transfection in just 1 day
  • Eliminates trial and error approach of synthetic siRNA

Application Features

  • Gene silencing
  • Target validation

References

  1. Morlighem, J.E. et al. (2007). Biotechniques. 42, 599-606.
  2. Yang, D. et al. (2002). Proc. Natl. Acad. Sci. USA. 99, 9942-9947.
  3. Calegari, F. et al. (2002). Proc. Natl. Acad. Sci. USA. 99, 14236-14240.
  4. Donze, O. and Picard, D. (2002). Nucleic Acids Res. 30, e46.

Protocols, Manuals & Usage

Protocols

  1. ShortCut RNase III Digestion Protocol (NEB #M0245)
  2. Purification of siRNA by Ethanol Precipitation from ShortCut digests (NEB #M0245)
  3. Transfection Guidelines for ShortCut® RNase III (NEB #M0245)

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 sold for research use only and not for resale in any form.  Commercial use of this product may require a license.  For license information, please contact the Licensing Office, New England Biolabs, Inc., 240 County Road, Ipswich, MA 01938.

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