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Restriction Endonucleases >
Restriction Endonucleases >
HpaI |
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Prices are in US dollars and valid only for US orders.
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Recognition Site:
isoschizomers | compatible ends | single letter code
Source:
A E. coli strain that carries the HpaI gene from Haemophilus parainfluenzae (ATCC 49669).
Reagents Supplied:
NEBuffer 4
Enzyme Properties
Activity in NEBuffers:
| NEBuffer 1: | | 25% | | NEBuffer 2: | | 50% | | NEBuffer 3: | | 10% | | 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 some combinations of overlapping |
Heat Inactivation:
No
Survival in a Reaction:
Minimum units to digest 1 µg of substrate DNA in 16 hours: 0.25 unit(s)
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 λ DNA in 1 hour at 37°C in a total reaction volume of 50 µl.
Concentration:
5,000 units/ml
Unit Assay Substrate:
λ DNA
Storage Conditions:
10 mM Tris-HCl
50 mM KCl
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 A
Notes
General notes:- Conditions of low ionic strength, high enzyme concentration, glycerol concentration > 5%, or pH > 8.0 may result in star activity.
FAQs
- When is star activity a problem for HpaI?
- What do we know about star activity with Hpal I?
- How do you recomend using HpaI?
- Does a certain salt or salt level inhibit activity of HpaI?
- Does spermidine increase activity of HpaI?
- What is Star Activity and how can it be avoided?
- How can this enzyme be inactivated?
- Is HpaI inhibited by dUTP incorporated at the site?
- Is HpaI active at 25°C?
- What is the molecular weight of HpaI?
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 50-fold overdigestion with HpaI, > 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 HpaI.
16-Hour Incubation:
A 50 μl reaction containing 1 μg of DNA
and 75 units of HpaI incubated for 16 hours at 37ºC
resulted in a DNA pattern free of detectable nuclease degradation as determined by agarose gel electrophoresis. However, fragments produced by noncanonical cleavage due to star activity may be observed with 10 units of enzyme in similar conditions.
Exonuclease Activity:
Incubation of a 50 μl reaction containing 80 units of HpaI with 1 μg of a mixture of single and double-stranded
[3H] E. coli DNA (205 cpm/μg) for 4 hours at 37ºC
released < 0.1% of the total radioactivity.
Endonuclease Activity:
Incubation of a 50 μl reaction containing 50 units of HpaI with 1 μg of
pUC19 DNA for 4 hours at 37ºC resulted
in < 10% conversion to RFII as determined by agarose gel electrophoresis.
Reagents Sold Separately
NEBuffer 4
Legal
Patents:
New England Biolabs, Inc.: U.S. Patent No. 5,298,404
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