Does BamHI have star activity?
After 50-fold overdigestion with BamHI, >95 % of the DNA fragments can be ligated and recut with this enzyme. Star activity: Conditions of low ionic strength, high enzyme concentration, glycerol concentration >5 % or pH >8.0 may result in star activity.
Which enzymes have star activity?
This altered specificity has been termed “star activity”. It has been suggested that star activity is a general property of restriction endonucleases (1) and that any restriction endonuclease will cleave noncanonical sites under certain extreme conditions, some of which are listed below.
What does BamHI do?
BamHI (from Bacillus amyloli) is a type II restriction endonuclease, having the capacity for recognizing short sequences (6 b.p.) of DNA and specifically cleaving them at a target site.
What is the star activity of restriction enzyme?
Star activity is the relaxation or alteration of the specificity of restriction enzyme mediated cleavage of DNA that can occur under reaction conditions that differ significantly from those optimal for the enzyme.
What causes star activity?
Generally, star activity (i.e., the enzyme cuts a sequence that is an imperfect match to its known target) is caused by nonoptimal reaction conditions.(1),(2) These include: High concentration of glycerol (>5% v/v). High ratio of enzyme units to micrograms of DNA.
What is BamHI compatible with?
Cleavage with two restriction endonucleases that produce compatible overhangs….Compatible Cohesive Ends and Generation of New Restriction Sites.
| Enzyme | Ligated To | Recleaved By |
|---|---|---|
| BamHI * (G/GATCC) | BstYI (G/GATCC) | AlwI, BamHI, BstYI, DpnII, NlaIV |
| BanI | ||
| (G/GTACC) | Acc65I | Acc65I, BanI, KpnI, NlaIV, RsaI |
| (G/GCGCC) | KasI | BanI, BsaHI, HaeII, HhaI, KasI, NarI, NlaIV |
How do you heat inactivate BamHI?
Incubate at 37°C for 1 h. Heat inactivation Up to 10 Units BamH I / µg DNA can be heat-inacti- vated by 15 min incubation at 65°C, higher enzyme concentrations can no more be completely inactivated under these conditions.