BsaI restriction endonuclease
BsaI, an IIs restriction endonuclease restriction endonuclease, is derived from arecombinant E. coli strain that carries the cloned and modified BsaI gene from Bacillus stearothermophilus.Which have the same specificity as native enzyme and reduced star activity. Its recognition sequence and cleavage sites are as follows:
5'······GGTCTC(N)············3'
3'······CCAGAG(NNNNN)···5'
Components
BsaI(20U/μL), 10xBsaI Buffer
Storage
Store at -25℃~-15℃, valid for 1 year (Avoid repeated freeze-thaw cycles).
Storage buffer
10mM Tris-HCl, 200mM NaCl, 1mM DTT 0.1mM EDTA, 200µg/ml Recombinant Albumin,50% Glycerol. (pH 7.4 @ 25°C).
Product features
High activity, Fast digestion;
1. Low star activity, ensuring accurate cutting like “scalpel”;
2. Without BSA and animal-origin free.
Methylation Sensitivity
Dam methylation: Not Sensitive;
Dcm methylation: Impaired by SomeCombinations of Overlapping;
CpG Methylation: Blocked by SomeCombinations of Overlapping.
Unit Definition
One unitisdefinedasthe amount of enzyme required to digest 1 µg of Internal control DNA in 1 hour at 37°C in a total reaction volume of 50 µL.
Quality Control
Protein Purity Assay (SDS-PAGE): The purity of Bsa I was ≥ 95% determined by SDS-PAGE analysis using Coomassie Blue detection.
RNase: 20 U of Bsa I with 1.6 μg MS2 RNA for 16hoursat37℃yieldsnodegradationas determined by agarose gel electrophoresis.
Non-Specific DNase Activity: 20 Uof BsaI with 1 μg PhiX174 DNA for 16 hours at 37 ℃ yields no excess DNA as determined by agarose gel electrophoresis.
Ligation and Recutting: After digestion of 1 μg
E.coli DNA: 2Uof VacciniavirusCapping Enzyme is screened for the presence of E. coli genomic DNA using TaqManqPCR with primers specific for the E. coli 16S rRNA locus. The E. coli genomic DNA contamination is ≤1 E. coli genome.
BacterialEndotoxin: LAL-test, accordingto ChinesePharmacopoeiaIV2020edition,gel limit test method, general rule (1143). Bacterial endotoxin content should be ≤10 EU/mg.
Reaction system and conditions
|
Component |
Volume |
|
BsaI（20 U/μL） |
1μL |
|
DNA |
1μg |
|
10 x BsaI Buffer |
5μL |
|
dd H2O |
Up to 50 μL |
Incubate at 37°C for 15-30 minutes. Heat inactivation：80°C for 20 min.
The recommendedreactionsystem and conditions can provide relatively good enzyme digestioneffect,whichisforreference only, pleaserefertotheexperimentalresultsfor details.
Applications
Restriction Enzyme DigestionFast Cloning.
Notes on use
1.The volume of enzyme ≤ 1 / 10 of the reaction volume.
2. Staractivitymayoccurwhenglycerol concentration is more than 5%.
3. Cleavageactivitymay occur when Substrate below the recommended ratio.
