Description
For molar concentration and related product information, please refer to:
In vitro Cas9 mediated digestion of DNA
1. Add the following components to a sterile, nuclease-free tube on ice:
|
Components |
Volume |
Final Concentration |
|
|
sgRNA (300 nM) |
2 µl |
~30 nM |
|
|
Cas9 Nuclease Protein (1000 nM) |
0.60 µl |
~30 nM |
|
|
10X Cas9 Nuclease Reaction Buffer |
2 μl |
1X |
|
|
Nuclease-free H2O |
12.4 μl |
– |
|
|
Pre-Incubate for 15 minutes at room temperature |
|||
|
Substrate DNA (30 nM) |
3 μl |
3 nM |
|
|
Total Volume |
20 μl |
– |
|
2. Collect all components by a brief centrifugation. Incubate the reaction at 37 °C for 1 hour.
3. Analyze fragments by running an agarose gel electrophoresis.
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Data Sheet
| Applications | Create double blunt end for recombination |
| Concentration | 1 µg/ µL, 6.26 µM |
| Formulation | Recombinant Cas9 wild type protein expressed in E. coli supplied in a buffer of 10 mM Tris-HCl (pH 7.4), 0.1 mM EDTA, 1 mM DTT, 300 mM NaCl, and 50% (v/v) Glycerol. |
| Host/Source | E.coli |
| Physical appearance | Sterile filtered colorless solution. |
| Reaction Conditions | Use 1X Cas9 Reaction Buffer and incubate at 37 °C. |
| Size | 50 µg, 313 pmol, with Reaction Buffer |
| Storage buffer | 10 mM Tris-HCl (pH 7.4), 0.1 mM EDTA, 1 mM DTT, 300 mM NaCl, and 50% (v/v) Glycerol. |
| Storage Conditions | -70°C. Avoid freeze thaw cycles. |
