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Restriction Endonucleases >
Restriction Endonucleases >
NheI |
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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 NheI gene from Neisseria mucosa heidelbergensis (ATCC 25999).
Reagents Supplied:
NEBuffer 2
BSA
Enzyme Properties
Activity in NEBuffers:
| NEBuffer 1: | | 100% | | NEBuffer 2: | | 100% | | 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:
65°C for 20 minutes
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 2
Supplemented with 100 μg/ml Bovine Serum Albumin
Incubate at
37°C.
1X NEBuffer 2:
10 mM Tris-HCl
50 mM NaCl
10 mM MgCl2
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 (HindIII digest) in 1 hour at 37°C in a total reaction volume of 50 µl.
Concentration:
10,000 units/ml and 50,000 units/ml
Unit Assay Substrate:
λ DNA (HindIII digest)
Storage Conditions:
10 mM Tris-HCl
250 mM NaCl
1 mM Dithiothreitol
0.1 mM EDTA
200 µg/ml BSA
50% Glycerol
0.15% Triton X-100
pH 7.4 @ 25°C
Storage Temperature:
-20°C
Diluent Compatibility:
Diluent C
Notes
General notes:- Cleaves to leave a 5´ CTAG extension which can be efficiently ligated to DNA fragments generated by AvrII, SpeI or XbaI.
- Inhibited by salt concentrations > 100 mM.
FAQs
- Is NheI affected by methylation?
- Are more units of NheI required to cut supercoiled DNA than lambda DNA?
- Is NheI inhibited by salt?
- Is NheI active at 25°C?
- Does NheI produce commonly used compatible ends?
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 100-fold overdigestion with NheI, > 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 NheI.
16-Hour Incubation:
A 50 μl reaction containing 1 μg of DNA
and 60 units of NheI 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 700 units of NheI 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 30 units of NheI with 1 μg of
ΦX174 RF I DNA for 4 hours at 37ºC resulted
in < 10% conversion to RFII as determined by agarose gel electrophoresis.
Reagents Sold Separately
NEBuffer 2
BSA
Companion Products
NheI-HF™
Legal
Patents:
New England Biolabs, Inc.: U.S. Patent No. 6,387,681
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