 |
|
 |
|
|
| Home >
Products >
Restriction Endonucleases >
Restriction Endonucleases >
TspMI |
|
|
|
 |
|
Prices are in US dollars and valid only for US orders.
|
Recognition Site:
isoschizomers | compatible ends | single letter code
Source:
Thermus species (P. Sharma)
Reagents Supplied:
NEBuffer 4 (10X)
Enzyme Properties
Activity in NEBuffers:
| NEBuffer 1: | | 50% | | NEBuffer 2: | | 75% | | NEBuffer 3: | | 50% | | NEBuffer 4: | | 100% |
When using a buffer other than the optimal (supplied) NEBuffer, it may be necessary to add more enzyme to achieve complete digestion.
Methylation Sensitivity:
| dam methylation: Not sensitive | | dcm methylation: Not sensitive | | CpG methylation: Blocked |
Activity at 37°C:
20%
Heat Inactivation:
No
Survival in a Reaction:
Minimum units to digest 1 µg of substrate DNA in 16 hours: 0.13 unit(s)
Reaction & Storage Conditions
Reaction Conditions:
1X NEBuffer 4
Incubate at
75°C.
1X NEBuffer 4:
20 mM Tris-acetate
50 mM potassium acetate
10 mM Magnesium Acetate
1 mM Dithiothreitol
pH 7.9 @ 25°C
Unit Definition:
One unit is defined as the amount of enzyme required to digest 1 µg of pUCAdeno plasmid DNA in 1 hour at 75°C in a total reaction volume of 50 µl.
Concentration:
5,000 units/ml
Unit Assay Substrate:
pUCAdeno plasmid DNA
Storage Conditions:
20 mM Tris-HCl
300 mM NaCl
1 mM DTT
1 mM EDTA
200 µg/ml BSA
50% Glycerol
0.1% Triton X-100
pH 8.0 @ 25°C
Storage Temperature:
-20°C
Diluent Compatibility:
Diluent B
Notes
General notes:- Conditions of low ionic strength, high enzyme concentration, glycerol concentration > 5%, or pH > 8.0 may result in star activity.
- TspMI is an isoschizomer of XmaI.
Temperature activity curve of activity:
37°C gives 20% optimal activity
65°C gives 100% optimal activity
75°C gives 100% optimal activity
80°C gives 200% optimal activity
Quality Control for Current Lot
Quality control values for a specific lot can be found on the datacard which accompanies each product.
Ligation and Re-cutting:
After a 5-fold overdigestion with TspMI, > 95%
of the DNA fragments can be ligated with T4 DNA Ligase (at a 5' termini concentration of 1-2 μM)
at 16ºC. Of these ligated fragments, > 95% can be recut with TspMI.
16-Hour Incubation:
A 50 μl reaction containing 1 μg of DNA
and 75 units of TspMI incubated for 16 hours at 75ºC
resulted in a DNA pattern free of detectable nuclease degradation as determined by agarose gel electrophoresis.
Exonuclease Activity:
Incubation of a 50 μl reaction containing 100 units of TspMI with 1 μg of a mixture of single and double-stranded
[3H] E. coli DNA (205 cpm/μg) for 4 hours at 75ºC
released < 0.14% of the total radioactivity.
Endonuclease Activity:
Incubation of a 50 μl reaction containing 5 units of TspMI with 1 μg of
ΦX174 RF I DNA for 4 hours at 75ºC resulted
in < 10% conversion to RFII as determined by agarose gel electrophoresis.
Reagents Sold Separately
NEBuffer 4
|
|
|
 |
|
|
