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| | | | DpnI | | | Receive a free DNA Ladder Sample with the purchase of this enzyme through 9/30/2010! | |
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R0176L |
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5,000 units
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20,000 units/ml |
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R0176S |
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1,000 units
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20,000 units/ml |
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Recognition Site:
| single letter code
Source:
An E. coli strain that carries the DpnI gene from Diplococcus pneumoniae G41 (S. Lacks).
Reagents Supplied:
NEBuffer 4 (10X)
Enzyme Properties
Activity in NEBuffers:
| NEBuffer 1: | | 100% | | NEBuffer 2: | | 100% | | NEBuffer 3: | | 75% | | 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 by overlapping | |
More information about: Methylation Sensitivity |
Heat Inactivation:
80°C for 20 minutes
Survival in a Reaction:
(+ + +) Suitable for an extended or overnight digestion. Enzyme is active > 8 hours.
More information about: Extended Digests with Restriction Enzymes
Reaction & Storage Conditions
Reaction Conditions:
1X NEBuffer 4
Incubate at
37°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 pBR322 DNA (dam methylated) in 1 hour at 37°C in a total reaction volume of 50 µl.
Concentration:
20,000 units/ml
Unit Assay Substrate:
pBR322 DNA (dam methylated)
Storage Conditions:
10 mM Tris-HCl
400 mM NaCl
1 mM Dithiothreitol
0.1 mM EDTA
200 µg/ml BSA
50% Glycerol
pH 7.4 @ 25°C
Storage Temperature:
-20°C
Diluent Compatibility:
Diluent B
Notes
General notes:- DpnI cleaves only when its recognition site is methylated. DNA purified from a dam+ strain will be a substrate for DpnI.
FAQs
- Will DpnI cleave hemimethylated DNA?
- What's the difference between DpnI, DpnII, MboI, and Sau3AI?
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 20-fold overdigestion with DpnI, approximately 25%
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 DpnI.
16-Hour Incubation:
A 50 μl reaction containing 1 μg of DNA
and 200 units of DpnI incubated for 16 hours at 37ºC
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 500 units of DpnI with 1 μg of a mixture of single and double-stranded
[3H] E. coli DNA (105 cpm/μg) for 4 hours at 37ºC
released < 0.1% of the total radioactivity.
Endonuclease Activity:
Incubation of a 50 μl reaction containing 100 units of DpnI with 1 μg of
ΦX174 RF I DNA for 4 hours at 37ºC resulted
in < 15% conversion to RFII as determined by agarose gel electrophoresis.
Reagents Sold Separately
NEBuffer 4
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