{"product_id":"neb-r0117s","title":"New England Biolabs, R0117S, AatII","description":"AatII has been reformulated with Recombinant Albumin (rAlbumin) beginning with Lot #10268404.  \u003cb\u003eRelated Categories\u003c\/b\u003e Restriction Endonucleases: A,, Time-Saver Qualified Restriction Enzymes  \u003cb\u003eApplications\u003c\/b\u003e Fast Cloning: Accelerate your cloning workflows with reagents from NEB,, Restriction Enzyme Digestion  \u003cb\u003eSpecification\u003c\/b\u003e \u003cb\u003eUnit Definition\u003c\/b\u003e One unit is defined as the amount of enzyme required to digest 1 µg of λ DNA in 1 hour at 37°C in a total reaction volume of 50 μl.  \u003cb\u003eReaction Conditions\u003c\/b\u003e 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)  \u003cb\u003eActivity in NEBuffers\u003c\/b\u003e NEBuffer™ r1.1: \u0026lt;10% NEBuffer™ r2.1: 50% NEBuffer™ r3.1: 50% rCutSmart™ Buffer: 100%  \u003cb\u003eDiluent Compatibility\u003c\/b\u003e Diluent B  \u003cb\u003eStorage Buffer\u003c\/b\u003e 300 mM NaCl 10 mM Tris-HCl 1 mM DTT 0.1 mM EDTA 500 µg\/ml Recombinant Albumin 50% Glycerol pH 7.4 @ 25°C  \u003cb\u003eHeat Inactivation\u003c\/b\u003e 80°C for 20 minutes  \u003cb\u003eMethylation Sensitivity\u003c\/b\u003e dam methylation: Not Sensitive dcm methylation: Not Sensitive CpG Methylation: Blocked  \u003cb\u003eFAQ\u003c\/b\u003e Q: Are there any factors that inhibit cleavage by AatII? A: AatII does not work at low ionic strength. It is also blocked by CpG methylation. Q: Is AatII active at 25°C? A: It is 25-50% active at 25°C. Q: Do I have to set-up digests with Time-Saver™ qualified enzymes for 5-15 minutes? Can I digest longer? A: NEB's Time-Saver™ enzymes have the benefit of working fast (5-15 minutes), but are also designed and qualified to withstand overnight digestions without degradation of DNA. Q: 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? A: The quality and cleanliness of the DNA are major factors that can affect enzyme activity. Restriction Enzymes active in rCutSmart® Buffer, but not as active in NEBuffer r2.1 and\/or r3.1 (our higher salt buffers), can be inhibited by salt in the reaction. DNA purification procedures that use spin columns can result in DNA solutions with significant levels of salt that can carry over into the reaction and inhibit enzyme activity. To prevent this, we recommend that the DNA solution added to the reaction be no more than 25% of the total reaction volume. This can be attained by adjusting the total reaction volume accordingly. Q: Are Protease Inhibitors acceptable for use with the Protein Deglycosylation Mix reaction? A: When a protein is denatured it is more susceptible to cleavage by proteases. For this reason a protease cocktail containing the following can be used with the Protein Deglycosylation Mix protocol:Aprotinin (10 μg\/ml final concentration)Benzamidine (1mM final concentration)Pepstatin (10 μg\/ml final concentration)Leupeptin (1μM final concentration)EGTA (1 mM final concentration)EDTA (1 mM final concentration)PMSF (1 mM final concentration)Make a 1000X concentrated stock of each inhibitor in water; except Pepstatin which should be dissolved in methanol.Note: PMSF has the ability to modify basic residues on glycoprotein substrates. Q: Which NEB restriction enzymes are supplied with Gel Loading Dye, Purple (6X)? A: All HF-restriction enzymes and most NEB non-HF restriction enzymes are supplied with Gel Loading Dye, Purple (6X). The non-HF restriction enzymes that come supplied with purple dye are: AatII AsiSI BsmBI-v2 EagI MboI NlaIII PvuI SmaI Acil AvrII BspHI EcoRI MboII NotI RsaI SpeI AfeI BamHI BsrGI EcoRV MluI NspI SacI StuI AflII BbsI ClaI Esp3I FseI MseI PacI SacII AgeI BglII DdeI HaeIII MspI PciI SalI XbaI AluI BsaI DpnI HindIII NcoI PmeI SapI XhoI ApaI BseYI DpnII HpaI NdeI PsiI-v2 SfaNI XmaI ApeKI BsiWI DraI KpnI NheI PstI SfiI XmnI AscI * All HF restriction enzymes are also supplied with Gel Loading Dye, Purple (6X) Q: Is this enzyme sensitive to dam, dcm or mammalian CpG methylation? A: Yes. This enzyme cannot cut recognition sites in mammalian gDNA that are blocked by CpG methylation. For up-to-date information about methylation sensitivities, please visit Dam-Dcm and CpG Methylation or REBASE. Q: Can you tell me more about the switch from BSA to Recombinant Albumin (rAlbumin) in NEBuffers? A: NEB is excited to announce that we have switched BSA-containing reaction buffers (NEBuffer 1.1, 2.1, 3.1 and CutSmart® Buffer) to Recombinant Albumin-containing buffers (NEBuffer r1.1, r2.1, r3.1 and rCutSmart™ Buffer). We are also in the process of moving all restriction enzyme formulations to contain rAlbumin. We feel that moving away from animal-containing products is a step in the right direction and are able to offer this enhancement at the same price. Q: Can Gel Loading Dye, Purple 6X (B7024) be stored in cold temperatures? A: The recommended storage temperature for Gel Loading Dye, Purple 6X, is room temperature (25°C). Commonly, the SDS may precipitate out of solution when the dye is stored on cold temperatures. This slight precipitate can be dissipated by letting the vial sit at room temperature for an hour, or warming gently for 10 minutes. Mix the vial a few times before use, to bring the solution back to homogeneity. ","brand":"New England Biolabs","offers":[{"title":"Default Title","offer_id":46835508871337,"sku":"R0117S","price":0.99,"currency_code":"USD","in_stock":true}],"url":"https:\/\/iright.com\/products\/neb-r0117s","provider":"Iright","version":"1.0","type":"link"}