SphI-HF^®

The following reagents are supplied with this product:

NEB #	Component Name	Component #	Stored at (°C)	Amount	Concentration
R3182S     -20       SphI-HF® R3182SVIAL -20 1 x 0.025 ml 20,000 units/ml   rCutSmart™ Buffer B6004SVIAL -20 1 x 1.25 ml 10 X   Gel Loading Dye, Purple (6X) B7024AVIAL 25 1 x 0.5 ml 6 X
R3182L     -20       SphI-HF® R3182LVIAL -20 1 x 0.125 ml 20,000 units/ml   rCutSmart™ Buffer B6004SVIAL -20 1 x 1.25 ml 10 X   Gel Loading Dye, Purple (6X) B7024AVIAL 25 1 x 0.5 ml 6 X
R3182M     -20       SphI-HF® R3182MVIAL -20 1 x 0.025 ml 100,000 units/ml   rCutSmart™ Buffer B6004SVIAL -20 1 x 1.25 ml 10 X   Gel Loading Dye, Purple (6X) B7024AVIAL 25 1 x 0.5 ml 6 X

Companion Products

• SphI

Materials Sold Separately

• rCutSmart™ Buffer
• Gel Loading Dye, Purple (6X)

1. Cleaves to leave a 3' CATG extension which can be efficiently ligated to DNA fragments generated by NlaIII.
2. Based on the stability of the enzyme in the reaction, incubations longer than 1 hr will not result in improved digestion, unless additional enzyme is added. Please refer to Restriction endonuclease survival in a reaction for more information regarding this topic.
3. Not sensitive to CpG, dcm, or dam methylation.

1. Optimizing Restriction Endonuclease Reactions
2. Protocol for Direct Digestion of gDNA during droplet digital PCR (ddPCR)
3. Restriction Digest Protocol
4. Double Digest Protocol with Standard Restriction Enzymes

• Activity at 37°C for Restriction Enzymes with Alternate Incubation Temperatures
• Activity of Restriction Enzymes in PCR Buffers
• Cleavage Close to the End of DNA Fragments
• Digestion of Agarose-Embedded DNA: Info for Specific Enzymes
• Double Digests
• Effects of CpG Methylation on Restriction Enzyme Cleavage
• Heat Inactivation
• NEBuffer Activity/Performance Chart with Restriction Enzymes
• Optimizing Restriction Endonuclease Reactions
• Restriction Endonucleases - Survival in a Reaction
• Restriction Enzyme Diluent Buffer Compatibility
• Restriction Enzyme Tips
• Restriction Enzymes for Droplet Digital PCR (ddPCR)
• Single Letter Codes
• Star Activity
• Traditional Cloning Quick Guide

• Alphabetized List of Recognition Sequences
• Cleavage of Supercoiled DNA
• Compatible Cohesive Ends and Generation of New Restriction Sites
• Dam-Dcm and CpG Methylation
• Frequencies of Restriction Sites
• Isoelectric Points (pI) for Restriction Enzymes
• Isoschizomers
• NEB Diluent and Buffer Table
• Time-Saver™ Qualified Enzymes
• Why Choose Recombinant Enzymes?

• Competitor Cross-Reference Tool
• DNA Sequences and Maps Tool

• Double Digest Finder
• Enzyme Finder
• NEBcutter™ v3.0
• NEBioCalculator®
• REBASE®

1. Is there a difference in cutting close to the ends between SphI-HF and SphI?
2. How does the level of star activity of SphI-HF compare to SphI?
3. Why does the HF version of the enzyme have a different recommended buffer than the wild type enzyme?
4. When should I choose the HF version of the enzyme?
5. When is star activity a concern?
6. When should I choose the High Fidelity (HF®) version of the enzyme?
7. Can the change in buffer preference of the HF enzyme be advantageous?
8. Will the HF enzyme produce elevated star activity when used in a buffer other than the one recommended?
9. What does it mean to be Time-Saver™ qualified?
10. How is the improvement in fidelity of HF restriction endonucleases quantitated?
11. What is the Fidelity Index (FI)?
12. What does HF® refer to following the name of a restriction enzyme?
13. Do I have to set-up digests with Time-Saver™ qualified enzymes for 5-15 minutes? Can I digest longer?
14. 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?
15. Can Gel Loading Dye, Purple 6X (B7024) be stored in cold temperatures?
16. Which NEB restriction enzymes are supplied with Gel Loading Dye, Purple (6X)?
17. Is this enzyme sensitive to dam, dcm or mammalian CpG methylation?
18. Can you tell me more about the switch from BSA to Recombinant Albumin (rAlbumin) in NEBuffers?
19. 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?

• Restriction Enzyme Troubleshooting Guide

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.

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]

• R3182M_v2
• R3182S_L_v2

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]

• R3182M_v1_0011212
• R3182M_v1_0011304
• R3182M_v1_0011310
• R3182S_L_v1_0011212
• R3182S_L_v1_0011304
• R3182S_L_v1_0011310
• R3182S_L_v1_0011403
• R3182M_v1_0011403
• R3182M_v1_0011409
• R3182S_L_v1_0011409
• R3182S_L_v1_0031504
• R3182M_v1_0031506
• R3182S_L_v1_0031509
• R3182M_v1_0031512
• R3182S_L_v1_0031512
• R3182S_L_v1_0031608
• R3182M_v1_0031611
• R3182S_L_v1_0031701
• R3182M_v1_0031706
• R3182S_L_v1_0031706
• R3182M_v1_0031711
• R3182S_L_v1_0031711
• R3182L_v1_10015094
• R3182S_v1_10015096
• R3182M_v1_10021245
• R3182S_v1_10021246
• R3182L_v1_10027907
• R3182S_v1_10027909
• R3182S_v1_10033681
• R3182S_v1_10043644
• R3182L_v1_10043643
• R3182M_v1_10043647
• R3182S_v1_10047908
• R3182L_v1_10054648
• R3182S_v1_10054650
• R3182S_v1_10058363
• R3182S_v1_10063206
• R3182M_v1_10065334
• R3182S_v1_10068132
• R3182L_v1_10070993
• R3182S_v1_10074544
• R3182M_v1_10080461
• R3182S_v1_10084581
• R3182M_v1_10090260
• R3182S_v1_10090261
• R3182S_v1_10091402
• R3182L_v1_10091405
• R3182M_v1_10091403
• R3182S_v1_10096862
• R3182S_v1_10107067
• R3182L_v1_10111300
• R3182M_v1_10107068
• R3182S_v1_10115829
• R3182S_v1_10118231
• R3182L_v1_10127887
• R3182S_v1_10127886
• R3182S_v1_10138393
• R3182L_v2_10141353
• R3182M_v2_10141351
• R3182S_v2_10141349
• R3182S_v2_10150017
• R3182M_v2_10155320
• R3182S_v2_10157902
• R3182L_v2_10168212
• R3182S_v2_10168208
• R3182S_v2_10177172
• R3182S_v2_10182975
• R3182S_v2_10192272
• R3182S_v2_10203051
• R3182L_v2_10204392
• R3182S_v2_10211692
• R3182M_v2_10213823
• R3182S_v2_10237548
• R3182L_v2_10240200
• R3182S_v2_10241262
• R3182M_v2_10242820
• R3182S_v2_10249269
• R3182L_v2_10264800
• R3182S_v2_10264943
• R3182S_v2_10273568
• R3182M_v2_10280860
• R3182S_v2_10287462
• R3182S_v2_10284663
• R3182L_v2_10305675
• R3182S_v2_10307157
• R3182S_v2_10324925
• R3182M_v2_10331533
• R3182L_v2_10338269

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

• SphI-HF^®
• rCutSmart™ Buffer
• Gel Loading Dye, Purple (6X)

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.