Sfi I

Description

Recognition site and hydrolysis position:

Source: An E.coli strain, that carries the cloned gene Sfi I from Streptomyces fimbriatus
Unit definition:
One unit of the enzyme is the amount required to hydrolyze 1 μg of T7 DNA in 1 hour at 50°C in a total reaction volume of 50 μl.
Assayed on:
T7 DNA
Optimal SE-buffer: SE-buffer G
Enzyme activity (%):

Storage conditions: 10 mM Tris-HCl (pH 7.5); 200 mM NaCl; 0.1 mM EDTA; 7 mM 2-mercaptoethanol; 200 μg/ml BSA; 50% glycerol. Store at -20°C.
Ligations:
After 10-fold overdigestion with enzyme more than 70% of the DNA fragments can be ligated and recut. In the presence of 10% PEG ligation is better.
Non-specific hydrolisis:
No nonspecific activity was detected after incubation of 1 μg of T7 DNA with 20 u.a. of enzyme for 16 hours at 50°C. Reagents Supplied with Enzyme 10 X SE-buffer G, BSA
Reagents Supplied with Enzyme:
Methylation sensitivity: Blocked by overlapping Dcm methylation (C^mCWGG): GGCCWGGNNGGCC.
Not blocked by overlapping Dcm methylation (C^mCWGG): GGCCNNNNNGGCCWGG.
Notes: Incubation at 37°C rusults in 75-100% activity.
To obtain 100% activity, BSA should be added to the 1 x reaction mix to a final concentration of 100 μg/ml.
Do not use BSA for long incubation.
References:
Qiang, B.-Q., Schildkraut, I. Nucleic Acids Res. 12: 4507-4515 (1984).
Dedkov V.S., Degtyarev S. Kh. The resctriction endonucleases detection in colonies of microorganisms Streptomyces and Nocardia // Translated from APPLIED BIOCHEMISTRY and MICROBIOLOGY (Russia) Vol 28, 1992, #. 2, 309- 313