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Restriction Endonucleases >
Restriction Endonucleases >
BsoBI |
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Recognition Site:
isoschizomers | compatible ends | single letter code
Source:
A E. coli strain that carries the BsoB I gene from Bacillus stearothermophilus JN2091 (D. Clark).
Reagents Supplied:
NEBuffer 2
Enzyme Properties
Activity in NEBuffers:
| NEBuffer 1: | | 10% | | NEBuffer 2: | | 100% | | NEBuffer 3: | | 100% | | NEBuffer 4: | | 50% |
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: Not sensitive |
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 2
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 in 1 hour at 37°C in a total reaction volume of 50 µl.
Concentration:
10,000 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.
- BsoBI is a thermophilic isoschizomer of AvaI.
- The recommended incubation temperature has been changed from 65°C to 37°C to minimize star activity.
FAQs
- What is Star Activity and how can it be avoided?
- Do degenerate recognition sites need to be palindromic?
- How can this enzyme be inactivated?
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 40-fold overdigestion with BsoBI, > 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 BsoBI.
16-Hour Incubation:
A 50 μl reaction containing 1 μg of λDNA
and 40 units of BsoBI 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 160 units of BsoBI 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.
BsoBI: DNA cocrystals (Mark van der Woerd and Alan Friedman, Purdue University; John Pelletier, Hong Ruan, Laurence Ettwiller and Shuang-yong Xu, New England Biolabs)
Reagents Sold Separately
NEBuffer 2
Legal
Patents:
New England Biolabs, Inc.: U.S. Patent No. 5,492,823
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