The Micrococcal Nuclease (MNase, EC 3.1.31.1, CAS 9013-53-0) produced by our company is a Ca²⁺-dependent endonuclease derived from Staphylococcus aureus, with a molecular weight of approximately 18.6 kDa and an optimal pH range of 7-10. It can cleave single-stranded, double-stranded, linear and circular DNA or RNA, generating mononucleotides and oligonucleotides with 3′-phosphate ends. It has higher cleavage efficiency for single-stranded nucleic acids, and its cleavage efficiency at the 5′ side of A/T/U is about 30 times that at G/C, showing "relative" non-specificity. MNase only attacks the DNA in the nucleosome linker region, while nucleosomal DNA is protected by histones, making it an essential tool for chromatin structure research. In Chromatin Immunoprecipitation (ChIP) assays, MNase can precisely digest chromatin into fragments of 200 bp or 1-5 nucleosomes under cross-linked (X-ChIP) or non-cross-linked (N-ChIP) conditions. It not only preserves protein-DNA interactions but also avoids random damage caused by sonication, and is widely used in the localization of transcription factors, histone modification and epigenetic regulation research. In addition, MNase can also be used to remove nucleic acids from cell lysates, reduce lysate viscosity and improve the efficiency of downstream experiments.
| Source | Recombinant expression in Escherichia coli |
|---|
| Molecular Weight | 18.6 kDa |
| Appearance | Sterile liquid |
| Storage Buffer | 5 mM Tris (pH 7.4), 50 mM NaCl, 1 mM EDTA, 50% Glycerol. 100× BSA is provided with this product for MNase dilution. |
| Enzyme Concentration | 2000 gel units/μl |
| Purity | ≥95% |
| Activity Definition | One Agarose Gel Unit is defined as the amount of enzyme required to digest 1 μg of Lambda DNA at 37℃ for 15 minutes, resulting in the accumulation of low molecular weight DNA fragments of ＜400 bp as determined by agarose gel electrophoresis. Another activity unit is Kunitz Unit. One Kunitz Unit is defined as the amount of enzyme required to release acid-soluble oligonucleotides that produce an absorbance increase of 1.0 OD at 260 nm at 37℃ for 30 minutes. 1000 Agarose Gel Units is approximately equal to 100 Kunitz Units. |
Components and Description
M1518341
| Component | 320KU
| 5×320KU
| Storage |
| M1518341A | MNase (2000 gel units/μl)
| 160μl
| 5×160μl
| -20℃. Avoid freeze/ Thaw cycle. |
| M1518341B | Reaction Buffer (10×) | 1.6ml | 5×1.6ml
| -20℃. Avoid freeze/ Thaw cycle. |
| M1518341C | BSA (100×) | 250μl | 5×250μl
| -20℃. Avoid freeze/ Thaw cycle. |
Product Applications
Degradation of nucleic acids in protein preparations, in vitro translation, reduction of cell lysate viscosity in non-mechanical cell lysis preparation, chromatin structure analysis, rapid RNA sequencing.
Product Advantages
When used in ChIP assays, MNase has the advantages of good sample integrity and mild digestion conditions, and is also commonly used for chromatin fragmentation in ChIP assays.
Usage Instructions
1. Dilution of MNase. If MNase dilution is required, to ensure enzyme stability and reduce its adsorption to containers, it is recommended to use 1× Reaction Buffer supplemented with 1× BSA as the dilution buffer for serial dilution. Example: Add 5 μl of MNase (2000 gel units/μl) to 45 μl of dilution buffer, pipette up and down thoroughly to mix, and 50 μl of MNase with a concentration of 200 gel units/μl is obtained. Then add 5 μl of this MNase (200 gel units/μl) to 45 μl of dilution buffer, pipette up and down thoroughly to mix, and 50 μl of MNase with a concentration of 20 gel units/μl is obtained. Perform serial dilution in this manner.
2. Digestion of nucleic acid or cell samples with MNase.
a. Add the corresponding reagents and samples according to the table below.
| Reagent | Volume (20 μl system) | Volume (50 μl system) | Final Concentration |
|---|
| Reaction Buffer (10×) | 2 μl | 5 μl | 1× |
| MNase* | 1 μl | 2.5 μl | 1~2000 gel units |
| BSA (100×)* | 0.2 μl | 0.5 μl | 1× |
| Sample | x μl | x μl | - |
| Water | (17-x) μl | (42-x) μl | - |
| Total Volume | 20 μl | 50 μl | - |
Note 1: MNase can be appropriately diluted with reference to Step 1, and the specific dilution multiple and usage amount need to be optimized by pre-experiments.
Note 2: If the sample contains no protein, add BSA to the reaction buffer to a final concentration of 1×; no BSA is required for cell samples.
b. Mix the reaction system gently and incubate at 37℃ for 15-30 minutes (or longer) until the nucleic acids are completely digested or the expected digestion effect is achieved.
c. Add an appropriate amount of 0.5 M EDTA (pH 8.0) to terminate the reaction, e.g., add 1 μl of 0.5 M EDTA (pH 8.0) to a 20 μl reaction system, and 2.5 μl of 0.5 M EDTA (pH 8.0) to a 50 μl reaction system.
3. Chromatin fragmentation in ChIP assays with MNase. Please refer to the literature of relevant products for the specific experiment.
Notes
1. This product contains 50% glycerol and will not freeze when stored at -20℃. Do not store at -80℃, as freeze-thaw cycles will reduce enzyme activity.
2. This product has high viscosity, ensure accurate sampling volume when pipetting; pipette up and down thoroughly to mix after sample addition to avoid bubble formation.
3. Ca²⁺ is a key catalytic cofactor for MNase, the reaction buffer must contain 1-5 mM Ca²⁺ to ensure the catalytic activity of the enzyme; the presence of metal ion chelators such as EDTA and EGTA in the reaction system will inhibit enzyme activity.
4. The salt ion concentration in the reaction system must be lower than 100 mM, as excessively high salt concentration will affect the activity of MNase.
5. If the sample contains no protein, add BSA to the reaction buffer to a final concentration of 1×.
6. This product is only for scientific research by professional personnel, not for clinical diagnosis or treatment, not for the production of food and pharmaceuticals, and not for storage in ordinary residential buildings.
7. For your safety and health, wear a lab coat and disposable gloves during operation.
