Recognition & cleavage sequence

Source: Bacillus globigii

Reaction conditions

1X EzBuffer III 37℃

1X FastCut Buffer, 37℃

FastCut Buffer

Enzynomics restriction enzyme can cut substrate DNA in 5-15 min with FastCut Buffer

1X EzBuffer III

50 mM Tris-HCl (pH 7.9 @ 25℃), 100 mM NaCl, 10 mM MgCl₂, 100 μg/ml BSA

Unit definition

One unit is defined as the amount of enzyme required for complete digestion of 1-μg bacteriophage λ at 37℃ for 1 hr in 50-μl reaction mixtures.

Storage

20 mM Tris-HCl (pH 7.5 @ 25℃), 200 mM NaCl, 0.1 mM EDTA, 10 mM 2-mercaptoethanol, 200 μg/ml BSA, 50% glycerol.

Dilution buffer: EzDiluent B

10 mM Tris-HCl (pH 7.4 @ 25℃), 300 mM NaCl, 0.1 mM EDTA, 1 mM dithiothreitol, 500 μg/ml BSA, 50% glycerol.

Heat Inactivation

Bgl I can be inactivated at 65℃ for 20 min.

Methylation sensitivity

dam methylation: Not sensitive

dcm methylation: Not sensitive

CpG methylation: Conditionally sensitive

Prolonged incubation

A minimum amount of enzyme required to digest 1-μg substrate DNA for 16 hr; 0.13 U.

Relative activity in EzBuffers

EzBuffer I: 75%

EzBuffer II: 75%

EzBuffer III: 100%

EzBuffer IV: 50%

FastCut Buffer: 100%

Note

Five-fold higher activity can be obtained at pH 9.5 than pH 7.5. Cleavage of supercoiled DNA requires more (10 fold) enzyme. The sequences of sticky ends produced by this enzyme are unique to each site, thus, this enzyme can be conveniently used for unique cloning strategies; replacement of a wild-type fragment with a mutant one in a plasmid. Cleavage of mammalian genomic DNA can be blocked by CpG methylation that partially overlaps its recognition sequence. It is sensitive to impurities in DNA and buffer condition.

●  Enzynomics_R043_Bgl I_manual

● Material Safety Data Sheet

- Enzynomics_MSDS_R043_Bgl I

● Quality control

Overdigestion assay

DNA digested for 16 hr in 50-μl reaction with 100 U of enzyme resulted in same the DNA band patterns as those obtained with 1 U of enzyme for 1 hr.

Endonuclease assay

Less than 5% of 1-μg of ΦX174 RFI is converted to RFII when the DNA is incubated with 50 U of enzyme at 37℃ for 4hr in 50-μl reaction. 

Ligation and recutting

More than 95% of DNA fragments (1 μg) digested with 50-fold excess enzyme can be ligated by T4 DNA ligase (400 U) at 16℃ for 16 hr. Of the ligation products, > 95% can be re-cut. 

Extreme pure (EP)

No detectable degradation of ³²P-end labeled single-stranded and double-stranded (5’-, 3’-overhang and blunt end) oligonucleotides occurred during incubation with 100 U of enzyme for 4 hr.