Attention Golden Gate Assembly users: BsaI-HFv2 (NEB #R3733) has been optimized for Golden Gate Assembly*. BsaI-HFv2 also works well for any protocol requiring DNA cutting by BsaI. This is the recommended enzyme for any purpose requiring digestion at the recognition sequence ...5′-GGTCTC(N1)/(N5)-3′...
*The requirements for digestion of DNA during Golden Gate Assembly are more demanding than what is encountered in traditional DNA cloning. For Golden Gate, the enzyme must function well in a buffer that might not be the same buffer as that normally provided for the restriction enzyme, and maintain activity for an extended time at elevated temperatures in a dynamic cutting/re-ligating reaction with competition for substrate binding between the endonuclease and ligase. Extensive testing has demonstrated superior performance of BsaI-HFv2 compared to both BsaI (NEB #R0535) and BsaI-HF (NEB #R3535) in this challenging Golden Gate assembly context where restriction enzyme efficiency and fidelity are critically important.
Having supplied restriction enzymes to the research community for over 40 years, NEB has earned the reputation of being the leader in enzyme technologies. Working continuously to be worthy of that distinction, NEB strives to develop enzymes of the highest purity and unparalleled quality.
- High Fidelity (HF®) version available (NEB #R3733) supplied with CutSmart® Buffer
- Used in Golden Gate Assembly
- Type IIS restriction enzymes recognize asymmetric DNA sequences and cleave outside of their recognition sequence
- 100% activity in CutSmart Buffer (over 210 enzymes are available in the same buffer) simplifying double digests
- Supplied with 1 vial of Gel Loading Dye, Purple (6X)
Restriction Enzymes in Golden Gate Assembly
Golden Gate Assembly Workflow
Reduce Star Activity with High-Fidelity Restriction Enzymes
Standard Protocol for Restriction Enzyme Digests
NEB TV Episode 15
CutSmart Restriction Enzyme Buffer
Restriction Enzyme Digest Protocol: Cutting Close to DNA End
Restriction Enzyme Digestion Problem: DNA Smear on Agarose Gel
Why is My Restriction Enzyme Not Cutting DNA?
BsaI has a High Fidelity version BsaI-HF®v2 (NEB #R3733).
High Fidelity (HF) Restriction Enzymes have 100% activity in CutSmart Buffer; single-buffer simplicity means more straightforward and streamlined sample processing. HF enzymes also exhibit dramatically reduced star activity. HF enzymes are all Time-Saver qualified and can therefore cut substrate DNA in 5-15 minutes with the flexibility to digest overnight without degradation to DNA. Engineered with performance in mind, HF restriction enzymes are fully active under a broader range of conditions, minimizing off-target products, while offering flexibility in experimental design.
Product SourceAn E. coli strain that carries the cloned BsaI gene from Bacillus stearothermophilus 6-55 (Z. Chen)
The following reagents are supplied with this product:
Store at (°C) Concentration CutSmart® Buffer -20 10 X
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- Restriction Endonucleases: B
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