BsaI

Bsa I , an IIs restriction endonuclease restriction endonuclease, is derived from a recombinant E.coli strain that carries the cloned and modified BsaI gene from Bacillus stearothermophilus. which have the same specificity as native enzymes and reduced star activity. Its recognition sequence and cleavage sites are as

follows:

5’······GGTCTC(N)···············3′

3’······CCAGAG(NNNNN)······5

Product component

Storage

Store at -20 ℃, valid for 1 year（ Avoid repeated freeze-thaw cycles）

Storage buffer

10 mM Tris-HCl, 200 mM NaCl, 1 mM DTT 0.1 mM EDTA, 200 µg/ml Recombinant Albumin,50% Glycerol. (pH 7.4 @ 25°C).

Unit Definition

One unit is defined as the 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 16 hours at 37 ℃ yields no degradation as determined by agarose gel electrophoresis.

Non-Specific DNase Activity: 20 U of Bsa I with 1 μg PhiX174 DNA for 16 hours at 37 ℃ yields no excess DNA as determined by agarose gel electrophoresis.

Star activity: 40 U of Bsa I with 1 μg λ DNA/HindIII for 16 hours at 37 ℃ results in a DNA pattern free of detectable nuclease degradation as determined by agarose gel electrophoresis.

Ligation and Recutting: After digestion of 1 μg λDNA/HindIII with 20 U BsaI, DNA fragments can be ligated with T4 DNA ligase at 16ºC. And these ligated fragments can be recut with BsaI.  

E.coli DNA: 2 U of Vaccinia virus Capping Enzyme is screened for the presence of E. coli genomic DNA using TaqMan qPCR with primers specific for the E. coli 16S rRNA locus. The E. coli genomic DNA contamination is≤1 E.coli genome.

Bacterial Endotoxin: LAL-test, according to Chinese Pharmacopoeia IV 2020 edition, gel limit test method, the general rule (1143). Bacterial endotoxin content should be ≤10 EU/mg.

Reaction system and conditions

Incubate at 37°C for 15-30 minutes Heat Inactivation：80°C for 20 min

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 Some Combinations of Overlapping;

CpG Methylation: Blocked by Some Combinations of Overlapping.

Notes on use

1. The volume of enzyme ≤ 1 / 10 of the reaction volume.

2. Star activity may occur when glycerol concentration is more than 5%.

3. Cleavage activity may occur when Substrate below the recommended ratio.

Restriction Enzyme Digestion， Fast Cloning.