|Source||Bacillus subtilis T30|
|Substrate specificity||The enzyme cleaves only C5-methylated DNA and doesn't cut unmodified DNA.|
The enzyme activity depends on number and position of methylated nucleotides in the recognition sequence.
|Assayed on||Double-stranded oligonucleotide|
5` GCTTGTACTTTA G(5mC)G G C ATTGATTCTCACCACG 3`
3` CGAACATGAAAT C G C(5mC)G TAACTAAGAGTGGTGC 5`
|Unit definition||One unit is defined as the amount of enzyme required to cleave 1 pmol of the double-stranded oligonucleotide of the above indicated structure in 1 hour at 37°C in a total reaction volume of 20 μl.|
|Optimal SE-buffer||SE-buffer BisI (10 mM Tris-HCl (pH 9,0 at 25°C); 10 mM MgCl2; 150 mM KCl; 1 mM DTT.)|
|Enzyme activity (%)|
|10 - 25||25 - 50||50 - 75||75 - 100||50 - 75||30|
|Reaction buffer||SE-buffer BisI,(10 mM Tris-HCl (pH 9,0 at 25°C); 10 mM MgCl2; 150 mM KCl; 1 mM DTT.)|
|Storage conditions||10 mM Tris-HCl (pH 7.5); 100 mM NaCl; 0.1 mM EDTA; 7 mM 2-mercaptoethanol; 50% glycerol; Store at -20°C.|
|Non-specific hydrolisis||No nonspecific activity was detected after incubation of 1 μg of pFsp4HI1 DNA (BamHI digest) with 1 unit of BisI for 16 hours at 37°C. The pFsp4HI1 plasmid carries a gene for Fsp4HI DNA-methyltransferase, which modifies DNA forming 5'-G(5mC)NGC-3'/3'-CGN(5mC)G-5'.
|Reagents Supplied with Enzyme||
10 X SE-buffer BisI|
|Methylation sensitivity||The enzyme cleaves only C5-methylated DNA and doesn't cut unmodified DNA.|
|Inactivation 20 minutes under||65oC|
|Notes||No detectable degradation of a double-stranded labeled oligonucleotide was observed after incubation with 1 unit of restriction endonuclease BisI for 4 hours at 37°C.|
|MSDS:||Download MSDS as PDF|
|References:||Degtyarev, S.Kh., Rechkunova, N.I., Grinev, A.A., Dedkov, V.S. Izv. Sib. Otd. Akad. Nauk SSSR 15: 25-26 (1989).|
Elena V. Chmuzh, Julia G. Kashirina, Julia E. Tomilova, Nina V. Mezentzeva, Vladimir S. Dedkov, Danila A. Gonchar, Murat A. Abdurashitov, Sergei Kh. Degtyarev Restriction endonuclease Bis I from Bacillus subtilis T30 recognizes methylated sequence 5’-G(m5C)↓NGC-3’ // Biotechnologia (russ.). 2005. N 3. P. 22-26
Degtyarev S.Kh., Chmuzh E.V., Abdurashitov M. A., Kashirina J.G., Dedkov V.S., Tomilova J.E., Mezentseva N.V., Gonchar D. A. Bacterial strain Bacillus subtilis - producer of BisI restriction endonuclease. // Russian Federation patent RU 2270859 C1 (2006).