R0102 FAQs for BsrBI, Restriction Endonucleases, Intl, NEB
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BsrBI FAQ

Q1: The NEB catalog has historically stated that activity in NEBuffer 4 is less than 100% yet this enzyme is now supplied with NEBuffer 4. What have you changed?
Q2: Has the conversion to NEBuffer 4 altered any of the properties of the restriction enzyme?
Q3: If I have an old tube of enzyme, what NEBuffer should I use?
Q4: Will the new enzyme work in the originally supplied NEBuffer?
Q5: Why is NEB switching this restriction enzyme to NEBuffer 4?
Q6: Is BsrBI affected by methylation?
Q7: Are there special considerations when ligating following a BsrBI digestion?
Q8: Can BsrBI be used at other reaction temperatures?

Q1: The NEB catalog has historically stated that activity in NEBuffer 4 is less than 100% yet this enzyme is now supplied with NEBuffer 4. What have you changed?

A1: All our restriction enzymes were re-assayed in both NEBuffer 4 vs. the previously supplied NEBuffer. In a few cases minor formulation changes were required to bring the activity to 100% in NEBuffer 4.


Q2: Has the conversion to NEBuffer 4 altered any of the properties of the restriction enzyme?

A2: In general the properties of the restriction enzyme remain the same although for some enzymes, some minor changes in heat inactivation, Time-Saver™ qualification, etc. were observed. All the updated information can be found on the supplied data card as well as at
www.neb.com.


Q3: If I have an old tube of enzyme, what NEBuffer should I use?

A3: Our restriction enzymes are color coded on the label for the appropriate NEBuffer and can either be used with the previously supplied NEBuffer or with NEBuffer 4. To ensure 100% cleavage in NEBuffer 4, additional units of enzyme and/or longer incubation time may be necessary.


Q4: Will the new enzyme work in the originally supplied NEBuffer?

A4: Yes it will. In all instances the restriction enzyme will have 100% activity in the originally supplied NEBuffer.


Q5: Why is NEB switching this restriction enzyme to NEBuffer 4?

A5: Our main goal is to simplify the NEBuffer system so that the majority of restriction enzymes are compatible in a single buffer. We now supply 162 restriction enzymes with NEBuffer 4 including all the new High Fidelity (HF) restriction enzymes.


Q6: Is BsrBI affected by methylation?

A6: Cleavage is slowed significantly by CG methylation.


Q7: Are there special considerations when ligating following a BsrBI digestion?

A7: When DNA is cut with BsrBI and then ligated, only 50% of these ligated sites regenerate BsrBI sites because its recognition sequence is non-palindromic. Ligated sites not recleavable by BsrBI are recleavable by either SacI or SacII.


Q8: Can BsrBI be used at other reaction temperatures?

A8: BsrBI is fully active from 37C to 50C.