Product Class: Restriction Endonuclease

BsaI-HF®v2
neb31 cloned at NEB recombinant engineered time saver dil_B 37 80 Heat CpG dcm

BsaI-HFv2 has been reformulated with Recombinant Albumin (rAlbumin) beginning with Lot #10156743. Learn more.

Learn about Ligase Fidelity and Push the Limits of Golden Gate Assembly (50+ fragments).

BsaI-HFv2 is NEB Type IIS restriction enzyme that has been optimized for Golden Gate Assembly and in protocols requiring DNNEB cutting by BsaI Works well for any protocol requiring DNNEB cutting by BsaI Recommended enzyme for any purpose requiring digestion at the recognition sequence 5-GGTCTC(N1)/(N5)-3
Isoschizomers | Single Letter Code

Product Introduction

High-Fidelity (HF®) restriction enzymes have the same specificity as native enzymes, but have been engineered for significantly reduced star activity and performance in a single buffer (rCutSmart™ Buffer).  All HF-restriction enzymes come with Gel Loading Dye, Purple (6X).  Enjoy the enhanced performance and added value of our engineered enzymes at the same price as the native enzyme:

  • Engineered for improved performance
  • Optimized for Golden Gate Assembly; maintains high activity levels in T4 DNA Ligase Buffer often used in this application
  • Type IIS restriction enzymes recognize asymmetric DNA sequences and cleave outside of their recognition sequence
  • 100% activity in rCutSmart Buffer
  • Time-Saver™ qualified for digestion in 5-15 minutes
  • Reduced star activity
  • NEB has developed convenient kits (using BsmBI-v2 and BsaI-HFv2) for performing Golden Gate Assembly.
  • Can be used with NEBridge® Ligase Master Mix for Golden Gate Assembly
  • Restriction Enzyme Cut Site: GGTCTC(1/5)

"I really, really love BsaI-HFv2. We are back to getting phenomenal GGA results again with very little background."

    -M.C, Principal Investigator, Davidson College, NC

Catalog # Size Concentration
R3733S 1000.0 units 20000 units/ml
R3733L 5000.0 units 20000 units/ml

Featured Videos

View Video Library
  • GoldenGateAssemblyWorkflow_720

    Golden Gate Assembly Workflow

  • GoldenGateDomestication_Anim_720

    Golden Gate Assembly Domestication Tutorial

  • Listen to DAD Informatics tools and NEB enzymes to enable complex one-pot Golden Gate Assemblies

    Listen to DAD Informatics tools and NEB enzymes to enable complex one-pot Golden Gate Assemblies

  • high complexity golden gate assembly targets

    Listen to DAD (Data-optimized Assembly Design) when constructing high-complexity Golden Gate Assembly targets

  • Golden-Gate-Assembly-Tool-Tutorial

    Golden Gate Assembly Tool Tutorial

  • KuceraBecky

    Restriction Enzymes in Golden Gate Assembly

  • Type IIS Restriction Enzymes function

    Function and Application of Type IIS Restriction Enzymes

Product Information

Description

BsaI-HF®v2 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′.

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 Source

An E. coli strain that carries the cloned and modified BsaI gene from Bacillus stearothermophilus 6-55 (Z. Chen)
This product is related to the following categories:
High-Fidelity (HF®) Restriction Endonucleases Products,
Restriction Endonucleases B Products,
Time-Saver Qualified Restriction Enzymes Products,
This product can be used in the following applications:
NEBridge® Golden Gate Assembly ,
Restriction Enzyme Digestion,
Fast Cloning: Accelerate your cloning workflows with reagents from NEB,
Restriction Enzyme Digestion

Reagents Supplied

Reagents Supplied

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration

Properties & Usage

Unit Definition

One unit is defined as the amount of enzyme required to digest 1 µg of pXba DNA in 1 hour at 37°C in a total reaction volume of 50 µl.

Reaction Conditions

1X rCutSmart™ Buffer
Incubate at 37°C

1X rCutSmart™ Buffer
50 mM Potassium Acetate
20 mM Tris-acetate
10 mM Magnesium Acetate
100 µg/ml Recombinant Albumin
(pH 7.9 @ 25°C)

Activity in NEBuffers

NEBuffer™ r1.1: 100%
NEBuffer™ r2.1: 100%
NEBuffer™ r3.1: 100%
rCutSmart™ Buffer: 100%

Diluent Compatibility

Storage Buffer

20 mM Tris-HCl
300 mM NaCl
0.1 mM TCEP
200 µg/ml Recombinant Albumin
50% Glycerol
pH 9 @ 25°C

Heat Inactivation

80°C for 20 minutes

Methylation Sensitivity

dam methylation: Not Sensitive
dcm methylation: Impaired by Some Combinations of Overlapping
CpG Methylation: Blocked by Some Combinations of Overlapping

Isoschizomers

Bso31I
BspTNI
Eco31I

Product Notes

  1. BsaI-HFv2 has the same specificity as BsaI (NEB #R0535), but it has been engineered for reduced star activity.
  2. Impaired by some combinations of overlapping dcm methylation and blocked by some combinations of overlapping CpG methylation.

Protocols, Manuals & Usage

Protocols

  1. Optimizing Restriction Endonuclease Reactions
  2. Golden Gate (24 Fragment) Assembly Protocol
  3. Restriction Digest Protocol
  4. Double Digest Protocol with Standard Restriction Enzymes
  5. 52-Fragment Golden Gate Assembly using BsaI-HF®v2 (NEB #E1601)
  6. Golden Gate Assembly Protocol for Using NEBridge® Golden Gate Assembly Kit (BsaI-HF®v2) (E1601)

Usage & Guidelines

Application Notes

Tools & Resources

Selection Charts

FAQs & Troubleshooting

FAQs

  1. When is star activity a concern?
  2. When should I choose the HF version of an enzyme?
  3. Can the change in buffer preference of the HF enzyme be advantageous?
  4. Will the HF enzyme produce elevated star activity when used in a buffer other than the one recommended?
  5. What does it mean to be Time-Saver™ qualified?
  6. Do I have to set-up digests with Time-Saver™ qualified enzymes for 5-15 minutes? Can I digest longer?
  7. I tested your restriction enzyme on the substrate DNA recommended by NEB, and it appears to be active, however it does not digest my DNA. What could be the reason?
  8. Which restriction enzymes are used in Golden Gate Assembly?
  9. Which restriction enzymes are used in GoldenBraid Assembly?
  10. Is Gel Loading Dye, Purple (6X) or Gel Loading Dye, Purple (6X), no SDS compatible with other DNA binding dyes such as SYBR® and GelRed™ during gel electrophoresis?
  11. Can Gel Loading Dye, Purple 6X (B7024) be stored in cold temperatures?
  12. How many nucleotides do I have to add adjacent to the RE recognition site in order to get efficient cutting?
  13. Which NEB restriction enzymes are supplied with Gel Loading Dye, Purple (6X)?
  14. What is the activity of the Type IIS restriction enzyme BsaI-HFv2 (NEB #R3733) in T4 DNA Ligase Buffer?
  15. Is this enzyme sensitive to dam, dcm or mammalian CpG methylation?
  16. Does NEB offer any BSA-free and/or animal origin-free restriction enzymes for linearization of plasmids for mRNA vaccine development?
  17. Can you tell me more about the switch from BSA to Recombinant Albumin (rAlbumin) in NEBuffers?
  18. Are certain restriction enzymes more active at higher incubation temperatures than what is recommended?

Troubleshooting

Quality, Safety & Legal

Quality Assurance Statement

Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.

Specification Change Notifications

BsaI-HFv2 has been reformulated with Recombinant Albumin (rAlbumin) beginning with Lot #10156743. All subsequent (higher number) lots will contain rAlbumin. For additional information, please visit at NEB Restriction Enzyme formulations with Recombinant Albumin (rAlbumin).

Specifications

The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]

Certificate Of Analysis

The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]

Safety DataSheets

The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

Legal and Disclaimers

Products and content are covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB). The use of trademark symbols does not necessarily indicate that the name is trademarked in the country where it is being read; it indicates where the content was originally developed. The use of this product may require the buyer to obtain additional third-party intellectual property rights for certain applications. For more information, please email busdev@neb.com.

This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.

Other Products You May Be Interested In