Home Nucleic Acid Purification ShortCut® RNase III

ShortCut® RNase III
Gray property icon for NEBU recombinant Gray property icon for incubation temperature 37° Gray property icon for heat inactivation no

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.
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Catalog # Cat # Concentration Conc Size
M0245S 2,000 units/ml 200 units
M0245L 2,000 units/ml 1,000 units
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  • 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

      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 fluorescently labelled 40 bp dsRNA to siRNA in 20 minutes at 37°C in a total reaction volume of 20 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.

    • Advantages and Features

      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

    • Related Products

      Companion Products

    • 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.

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