| Name | | BstDE I |   |
| | Cat. # | E227T | E228T | | Number of reactions | 50 | 250 | | Volume, μl | 50 | 250 |
| Recognition site | C↑TNAG
GANT↓C | | Source | Bacillus stearothermophilus DE | | Assayed on | Lambda DNA, plasmid DNA | | Description | Turbo BstDE I is used for a short time (10 min) of 1 μg DNA digestion in universal (ROSE) SE-Buffer in 50 μl of reaction mixture.
Reaction Original SibEnzyme (ROSE) Buffer is a specially designed universal reaction buffer for the most Restriction Endonucleases. ROSE Buffer is perfect for DNA cleavage with SE Turbo Restriction Endonucleases and for double digestion.
Applications:
- Fast DNA analysis
- Fast preparation of vectors for cloning
- Double digestion
| | Reaction buffer | SE-buffer ROSE | | Optimal temperature | 60oC | | 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. | | Ligation | After digestion with 1 μl of Turbo BstDE I, approximately 90% of the DNA fragments can be ligated with high-activity T4 DNA Ligase and recut. | | Non-specific hydrolisis | No detectable degradation of a single-stranded and double-stranded oligonucleotide was observed after incubation with 1 μl of restriction endonuclease for 3 hours.
| | Reagents Supplied with Enzyme |
10 X SE-buffer ROSE | | Methylation sensitivity | not tested | | Inactivation 20 minutes under | 80oC | | Notes | Enzyme Properties:
1 μl of Turbo BstDE I cuts 1 μg of DNA in 1x SE-Buffer ROSE in 10 min (assayed on Lambda and plasmid DNA). A short time of DNA digestion requires high quality purification of DNA sample (PCR fragments should be purified after amplification).
Please note that supercoiled plasmid DNA and PCR fragments may have varying rates of cleavage and sometimes need more time to be completely digested.
Standard protocol of Turbo reaction:
20 μl of the reaction volume:
10x SE-Buffer ROSE - 2 μl
DNA - 0.2-1 μg
Nuclease-free water - to 20 μl
+ 1 μl of Turbo BstDE I
Mix by pipette tip carefully.
Incubate at 60°C for 10 min.
| | References: | Shinkarenko, N.M., Shevchenko, A.V., Dedkov, V.S., Abdurashitov, M.A., Degtyarev, S.K. Unpublished observations (1995).
 Murat A Abdurashitov, Victor N Tomilov, Valery A Chernukhin, S. Kh. Degtyarev A physical map of human Alu repeats cleavage by restriction endonucleases // BMC Genomics 2008, 9:305
Abdurashitov M.A., Tomilov V.N., Chernukhin V.A., Gonchar D. A., Degtyarev S. Kh. Comparative analysis of human chromosomal DNA digestion with restriction endonucleases in vitro and in silico. // "Medical genetics" V.6, No 8, pp 29-36, 2007
V.A. Chernukhin, M.A. Abdurashitov, V.N. Tomilov, D.A. Gonchar, S.Kh. Degtyarev Comparative restriction enzymes analysis of rat chromosomal DNA in vitro and in silico // "Ovchinnikov bulletin of biotechnology and physical and chemical biology" V.2, No 3, pp 39-46, 2006
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