USER® Enzyme
Product informationCode | Name | Size | Quantity | Price | |
---|---|---|---|---|---|
M5505S |
USER Enzyme, recombinant |
50 units ( 1000 units/ml ) | - | Unavailable in your region | |
M5505L |
USER Enzyme, recombinant |
250 units ( 1000 units/ml ) | - | Unavailable in your region |
USER® Enzyme
USER Enzyme has been reformulated with Recombinant Albumin (rAlbumin) beginning with Lot #10233215. Thermolabile USER II Enzyme (NEB #M5508) and Thermostable USER III Enzyme (NEB #M5509) are available.
Product Introduction
USER (Uracil-Specific Excision Reagent) Enzyme (1) generates a single nucleotide gap at the location of a uracil residue.
Applications of USER include:
- Directional RNA Seq
- NEBNext adaptor cleavage
- USER cloning
Figure 1: USER Enzyme
Catalog # | Size | Concentration |
---|---|---|
M5505S | 50.0 units | 1000 units/ml |
M5505L | 250.0 units | 1000 units/ml |
- Product Information
- Protocols, Manuals & Usage
- Tools & Resources
- FAQs & Troubleshooting
- Citations & Technical Literature
- Quality, Safety & Legal
- Other Products You May Be Interested In
Product Information
Description
USER (Uracil-Specific Excision Reagent) Enzyme (1) generates a single nucleotide gap at the location of a uracil. USER Enzyme is a mixture of Uracil DNA glycosylase (UDG) and the DNA glycosylase-lyase Endonuclease VIII. UDG catalyses the excision of a uracil base, forming an abasic (apyrimidinic) site while leaving the phosphodiester backbone intact (2,3). The lyase activity of Endonuclease VIII breaks the phosphodiester backbone at the 3´ and 5´ sides of the abasic site so that base-free deoxyribose is released (4,5).To help select the best DNA assembly method for your needs, please use our Synthetic Biology/DNA Assembly Selection Chart.
Product Source
The two component proteins are purified separately from E. coli K-12 strains containing plasmids encoding Endonuclease VIII and Uracil-DNA Glycosylase.- This product is related to the following categories:
- DNA Repair Enzymes and Structure-specific Endonucleases Products,
- This product can be used in the following applications:
- Applications of USER® and Thermolabile USER II Enzymes,
- USER® Cloning,
- DNA Assembly and Cloning, High-throughput cloning and automation solutions
Reagents Supplied
Reagents Supplied
The following reagents are supplied with this product:
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Properties & Usage
Unit Definition
One unit is defined as the amount of enzyme required to nick 10 pmol of a 34 mer oligonucleotide duplex containing a single uracil base, in 15 minutes at 37°C in a total reaction volume of 10 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)
Storage Buffer
10 mM Tris-HCl
5 mM NaCl
1 mM DTT
0.1 mM EDTA
175 µg/ml Recombinant Albumin
50% Glycerol
50 mM KCl
pH 7.4 @ 25°C
Heat Inactivation
NoRelated Products
Companion Products
Product Notes
- USER Enzyme is active in all commercial PCR buffers tested. It also has 100% activity in TE (10 mM Tris-HCl pH 8.0, 0.1 mM EDTA).
- Incubate at 37°C.
References
- Bitinaite, J. unpublished observations.
- Lindhal, T., Ljungquist, S., Siegert, W., Nyberg, B. and Sperens, B. (1977). J. Biol. Chem.. 252, 3286-3294.
- Lindhal, T. (1982). Annu. Rev. Biochem.. 51, 61-64.
- Melamede, R.J., Hatahet, Z., Kow, Y.W., Ide, H. and Wallace, S.S. (1994). Biochemistry. 33, 1255-1264.
- Jiang, D., Hatahet, Z., Melamede, R.J., Kow, Y.W. and Wallace, S.S. (1997). J. Biol. Chem.. 272, 32230-32239.
Protocols, Manuals & Usage
Protocols
Usage & Guidelines
Tools & Resources
Selection Charts
FAQs & Troubleshooting
FAQs
- Will the USER Enzyme work in rCutSmart buffer?
- Where can I find information or a protocol for cloning with USER® Enzyme?
- During an NEBNext DNA library prep, which enzyme cleaves NEBNext DNA loop adaptors and uracil-containing single stranded loops?
- Can you tell me more about the switch from BSA to Recombinant Albumin (rAlbumin) in NEBuffers?
- What is the difference between the suite of USER® enzymes (#M5505, #M5508, #M5509) and Thermostable Endonuclease Q (#M0701)?
Citations & Technical Literature
Citations
Additional Citations
- Cavaleiro, A.M., Kim, S.H., Seppälä, S., Nielsen, M.T., Nørholm, M.H.H. (2015) Accurate DNA Assembly and Genome Engineering with Optimied Uracil Excision Cloning ACS Synth Biol; 4 (9), 1042-1046. DOI: 10.1021/acssynbio.5b00113
- Salomonsen B., Mortensen U.H., Halkier B.A. (2014) USER-Derived Cloning Methods and Their Primer Design In: Valla S., Lale R. (eds) DNA Cloning and Assembly Methods Methods in Molecular Biology (Methods and Protocols); 116, 59-72. DOI: 10.1007/978-1-62703-764-8_5
- Wang S, Li W, Wang S, Hu B (2014) Rapid and Efficient Assembly of Transcription Activator-Like Effector Genes by USER Cloning J Genet Genomics; 41(6), 339-47. PubMedID: 24976123, DOI: 10.1016/j.jgg.2014.05.002
- Lund, A.M., Kildegaard, H.F., Petersen, M.B.K., Rank, J., Hansen, B.G., Andersen, M.R., et al. (2014) A Versatile System for USER Cloning-Based Assembly of Expression Vectors for Mammalian Cell Engineering PLoS One; 9 (5), e96693. DOI: 10.1371/journal.pone.0096693
- Lund AM, Kildegaard HF, Petersen MB, Rank J, Hansen BG, Andersen MR, Mortensen UH (2014) A versatile system for USER cloning-based assembly of expression vectors for mammalian cell engineering PLoS One; 9(5), e96693. PubMedID: 24879460, DOI: 10.1371/journal.pone.0096693
- Villiers BR, Stein V, Hollfelder F (2010) USER friendly DNA recombination (USERec): a simple and flexible near homology-independent method for gene library construction Protein Eng Des Sel; 23(1), 1-8. PubMedID: 19897542, DOI: 10.1093/protein/gzp063
- Nour-Eldin H.H., Geu-Flores F., Halkier B.A. (2010) USER Cloning and USER Fusion: The Ideal Cloning Techniques for Small and Big Laboratories In: Fett-Neto A. (eds) Plant Secondary Metabolism Engineering. Methods in Molecular Biology (Methods and Protocols); 643, 185-200. DOI: 10.1007/978-1-60761-723-5_13
Publications
- Thuronyi, B.W., Koblan, L.W., Levy, J.M. et al (2019) Continuous evolution of base editors with expanded target compatibility and improved activity Nat Biotechnol; 37, 1070-1079.. DOI: 10.1038/s41587-019-0193-0
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
USER Enzyme has been reformulated with Recombinant Albumin (rAlbumin) beginning with Lot #10233215. All subsequent (higher number) lots will contain 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]- M5505S_L_v1_0161711
- M5505S_L_v1_0161804
- M5505L_v1_10014935
- M5505L_v1_10021299
- M5505S_v1_10022311
- M5505L_v1_10026308
- M5505L_v1_10030343
- M5505L_v1_10032161
- M5505L_v1_10036514
- M5505S_v1_10036511
- M5505L_v1_10043461
- M5505L_v1_10050203
- M5505L_v1_10054976
- M5505S_v1_10055998
- M5505L_v1_10059016
- M5505L_v1_10063251
- M5505S_v1_10064310
- M5505L_v1_10068456
- M5505L_v1_10071829
- M5505L_v1_10081766
- M5505S_v1_10083543
- M5505L_v1_10085915
- M5505L_v1_10091116
- M5505S_v1_10091172
- M5505L_v1_10091565
- M5505S_v1_10092240
- M5505L_v1_10093554
- M5505S_v1_10098884
- M5505L_v1_10103303
- M5505L_v1_10107925
- M5505S_v1_10111504
- M5505L_v1_10114217
- M5505L_v1_10117538
- M5505L_v1_10126652
- M5505L_v1_10131502
- M5505L_v1_10136013
- M5505L_v1_10140665
- M5505S_v1_10136014
- M5505L_v1_10147205
- M5505L_v1_10153113
- M5505S_v1_10159204
- M5505L_v1_10159206
- M5505L_v1_10166598
- M5505L_v1_10171164
- M5505S_v1_10175983
- M5505L_v1_10176565
- M5505L_v1_10182267
- M5505L_v1_10190503
- M5505L_v1_10195920
- M5505S_v1_10206718
- M5505L_v1_10205073
- M5505L_v1_10214502
- M5505L_v1_10229458
- M5505L_v2_10233218
- M5505S_v2_10233217
- M5505L_v2_10238792
- M5505S_v2_10246077
- M5505L_v2_10246078
- M5505L_v2_10253492
- M5505S_v2_10253493
- M5505L_v2_10263599
Safety DataSheets
The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.USER® Enzyme
rCutSmart™ Buffer
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
The supporting documents available for this product can be downloaded below.