Product info: Bsp19 I


Name
Bsp19 Inew package    
Cat. #E047TE048T
Number of reactions50250
Volume, μl50250

Recognition site
CCATGG
GGTACC
SourceBacillus species 19
Assayed onLambda DNA, plasmid DNA
DescriptionDescription: Turbo Bsp19 I is used for a short time (10 min) of 1 μg DNA digestion in SE-Buffer W+ in 50 μl of reaction mixture.
Applications:
- Fast DNA analysis
- Fast preparation of vectors for cloning
- Double digestion
Reaction bufferSE-buffer 2W+ (20 mM Tris-HCl (pH 8,5 at 25°C); 10 mM MgCl2; 200 mM NaCl; 1 mM DTT ; 100 μg/ml BSA)
Optimal temperature
37oC
Storage conditions10 mM Tris-HCl (pH 7.5); 50 mM KCl; 0.1 mM EDTA; 7 mM 2-mercaptoethanol; 200 μg/ml BSA; 50% glycerol. Store at -20°C.
LigationAfter digestion with 1 μl of Turbo Bsp19 I, approximately 90% of the DNA fragments can be ligated with high-activity T4 DNA Ligase and recut.
Non-specific hydrolisisNo 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 2W+
Methylation sensitivityBsp19I cuts hemimethylated site
5`-(5mC)CATGG-3`/3`-GGTACC-5`
and doesn't cut methylated sites
5`-(5mC)CATGG-3`/3`-GGTAC(5mC)-5` and
5`-(4mC)CATGG-3`/3`-GGTAC(4mC)-5`.
Inactivation 20 minutes under
65oC
NotesEnzyme Properties:
1 μl of Turbo Bsp19 I cuts 1 μg of DNA in 1x SE-Buffer W+ in 10 min (assayed on Lambda DNA 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 W+ - 2 μl
DNA - 0.2-1 μg
Nuclease-free water - to 20 μl
+ 1 μl of Turbo Bsp19 I
Mix by pipette tip carefully.
Incubate at 37°C for 10 min.
References:Repin, V.E., Rechkunova, N.I., Degtyarev, S.Kh. Unpublished observations. Repin, V.E., Lebedev, L.R., Andreeva, I.S., Puchkova, L.I., Zernov, Y.P., Serov, G.D., Tereshchenko, T.A., Aphinogenova, G.N., Pustoshilova, N.M. Biotekhnologiya 0: 18-27 (1998).
 Chernukhin V.A, Abdurashitov M.A., Tomilov V.N., Gonchar D.A., Degtyarev S.Kh. Comparative analysis of mouse chromosomal DNA digestion with restriction endonucleases in vitro and in silico // "Ovchinnikov bulletin of biotechnology and physical and chemical biology" V.3, No 4, pp 19-27, 2007