To view the entire NEB catalog, visit their website here. Note: Also available as a FastDigest enzyme. We are excited to announce that all reaction buffers are now BSA-free. Roche There is no buffer available in which both enzymes exhibit 100 activity As these two enzymes are not compatible, Roche recommends to perform a sequential digestion to get the job done. each enzyme works at regular 1-fold concentration to obtain 100 cleavage. See Reaction Conditions for Restriction Enzymes for a table of enzyme activity, conditions for double digestion, and heat inactivation for this and other restriction enzymes. XhoI has been reformulated with Recombinant Albumin (rAlbumin) beginning with Lot 10161948. 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). KpnI-HF/NotI-HF both exhibit 100 activities in NEBuffer 4, i.e. ![]() ![]() The items we carry in stock are listed in the table below as well as in the NEB UMaine Freezer Program website. Thermo Scientific NotI restriction enzyme recognizes GCGGCCGC sites and cuts best at 37☌ in O buffer (Isoschizomers: CciNI). To avoid these, follow the recommended guidelines for storage and reactions, and always check for the efficacy of digestion along with purification of digested products on an agarose gel.The University of Maine Freezer Program receives a 15% discount on all products available in the New England Biolabs (NEB) catalog. Individual CBSs were PCR amplified from mouse. Star activity (or off-target cleavage) and incomplete cleavage are potential challenges which may occur due to suboptimal enzymatic conditions or inappropriate enzyme storage. A high fidelity restriction endonuclease that recognizes the sequence GCGGCCGC. The donor vector was then digested with the enzyme cocktail of NotI-HF (NEB, R3642S), SbfI-HF (NEB, R3189S) and rSAP (NEB, M0371S) for 4 h at 37 C. For complete digestion, make sure that the enzyme volume is 1/10th of the total reaction volume, the optimal temperature is constantly maintained throughout the reaction, the total reaction time is appropriately calculated based on the amount of DNA to be digested, appropriate buffers should be used to ensure maximal enzymatic activity, and in case of a double digest, make sure that the two restriction sites are far enough so that the activity of one enzyme cannot interfere with the activity of the other. NotI has a High Fidelity version NotI-HF ® ( NEB R3189 ). Always use a control DNA digestion with the enzyme to ensure adequate activity (to avoid interference due to high glycerol in the enzyme). The enzyme should always be stored at -20C and multiple freeze-thaw cycles should be avoided in order to maintain optimal activity. The most common challenges with restriction digest include- 1. Our high quality reagents are available for every workflow, including popular DNA assembly methods such as NEBuilder HiFi DNA Assembly and NEBridge Golden Gate Assembly. ![]() HF enzymes also exhibit dramatically reduced star activity. Whether you are performing your first cloning experiment or constructing multi-fragment DNA assemblies, NEB has the solution for you. High Fidelity (HF) Restriction Enzymes have 100 activity in rCutSmart Buffer single-buffer simplicity means more straightforward and streamlined sample processing. NEB began switching our BSA-containing reaction buffers in April 2021 to buffers containing Recombinant Albumin (rAlbumin) for. The four most common types of restriction enzymes include: Type I (cleaves at sites remote from a recognition site), Type II (cleaves within or at short specific distances from a recognition site), Type III (cleave at sites a short distance from a recognition site), and Type IV (targets modified DNA- methylated, hydroxymethylated and glucosyl-hydroxymethylated DNA). NotI has a High Fidelity version NotI-HF ( NEB R3189 ). SpeI-HF has been reformulated, and now also includes Recombinant Albumin (rAlbumin), beginning with Lot 10192521. Restriction Enzymes NotI A restriction enzyme or restriction endonuclease is defined as a protein that recognizes a specific, short nucleotide sequence and cuts the DNA only at or near that site, known as restriction site or target sequence.
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