Acu I

Description

Recognition site and hydrolysis position:

Source: An. E.coli strain that carries the cloned Acu I gene from Acinetobacter calcoaceticus SRW4
Unit definition:
One unit of the enzyme is the amount required to hydrolyze 1 μg of Lambda DNA in 1 hour at 37°C in a total reaction volume of 50 μl.
Assayed on:
Lambda DNA
Optimal SE-buffer: SE-buffer Y + SAM + BSA
Enzyme activity (%):

Storage conditions: 10 mM Tris-HCl (pH 7.5); 100 mM NaCl; 0.1 mM EDTA; 1 mM DTT; 200 μg/ml BSA; 50% glycerol; Store at -20°C.
Ligations:
After 2-fold overdigestion with enzyme about 80% of the DNA fragments can be ligated. Of these, 80% can be recut.
Non-specific hydrolisis:
No nonspecific activity was detected after incubation of 1 μg of Lambda DNA with 1 u.a. of enzyme for 16 hours at 37°C. Reagents Supplied with Enzyme 10 X SE-buffer Y, BSA, SAM
Reagents Supplied with Enzyme:
Methylation sensitivity: not tested
Notes: High enzyme concentration may result in star activity.
To obtain 100% activity, BSA should be added to the 1x reaction mix to a final concentration of 100 μg/ml and SAM should be added to a final concentration 10 μM. It is possible to use a final SAM concentration of 10-64 μM. In this case 32 mM SAM solution should be diluted in 3200-500 times, correspondingly. For the small volume of reaction mix (10-20 μl) the stock SAM solution may be previously diluted up to 1 mM (in 32 times) with 5 mM solution of H₂SO₄ or sterile water and stored at -20°C.
Do not use BSA for long incubation.
References:
Degtyarev, S.Kh., Kileva, E.V., Dedkov, V.S. Unpublished observations (2001).