Not I
* Size:

Not I Technical BrochureTechnical Brochure

NotI is a restriction enzyme purified from Nocardia otitidis-caviarum.


Unit substrate: Adenovirus-2 DNA.


Unit calculation assay conditions: 100 mM NaCl, 50 mM Tris-HCl (pH 7.9 @ 25°C), 5 mM MgCl2, 1 mM dithiothreitol, 100 μg/ml BSA. Incubate at 37°C.


Absence of contaminants: 80 units of Not I do not produce any unspecific cleavage products after 16 hrs incubation with 1 μg of Adeno-2 DNA at 37°C. After 30-fold overdigestion with Not I, greater than 98% of the DNA fragments can be ligated and recut with this enzyme.


Storage buffer: 500 mM KCl, 10 mM Tris-HCl (pH 7.4), 0.1 mM EDTA, 1mM dithiothreitol, 0.1% Triton X-100, 500 μg/ml BSA and 50% glycerol. Store at -20oC.


Heat inactivation: 65oC for 20 minutes.


Methylation Sensitivity:

dam methylation: Not sensitive

dcm methylation: Not sensitive

CpG methylation: Blocked

Note: Supercoiled plasmids may require up to 5-fold more Not I for complete digestion than linear DNAs.


Percent Activity in MINOTECH Buffers
 L SH 
 <10 25-50  75-100  75  50  50 
General reaction mixture:
10U  1μl
10x UNotI buffer*  2μl 
DNA substrate  <1μg
Sterile ultrapure water  Up to 20 μl
Incubate for 15 min at 37oC  
*We recommend the addition of BSA to a final concentration of 100 μg/ml. 


Reagents supplied: 10x UNotI  buffer