{"product_id":"neb-r0544s","title":"New England Biolabs, R0544S, KasI","description":"KasI has been reformulated with Recombinant Albumin (rAlbumin) beginning with Lot #10211225.  \u003cb\u003eRelated Categories\u003c\/b\u003e Restriction Endonucleases H M  \u003cb\u003eApplications\u003c\/b\u003e 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 pBR322 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: 50% NEBuffer™ r2.1: 100% NEBuffer™ r3.1: 50% rCutSmart™ Buffer: 100%  \u003cb\u003eDiluent Compatibility\u003c\/b\u003e Diluent B  \u003cb\u003eStorage Buffer\u003c\/b\u003e 100 mM KCl 10 mM Tris-HCl 1 mM DTT 0.1 mM EDTA 50% Glycerol 200 µg\/ml Recombinant Albumin pH 7 @ 25°C  \u003cb\u003eHeat Inactivation\u003c\/b\u003e 65°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: Is KasI affected by methylation? A: KasI is blocked by CpG methylation. For up-to-date information about methylation sensitivities, please visit Dam-Dcm and CpG Methylation or REBASE. Q: Is extended digestion of KasI recommended? A: Extended digestion is not beneficial because the enzyme is not stable in reaction. Q: Are more units of KasI required to cut supercoiled DNA than lambda DNA? A: Yes, 4 untis are required to cleave 1 μg of litmus DNA. Q: Is activity loss of KasI seen in 6 months or less? A: Yes, loss can be seen after 6 months storage at -20°C. KasI should be stored at –80°C; to avoid multiple freeze-thaws please aliquot the enzyme and store at –80°C. 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: What isoschizomers are there? A: NEB sells two neoschizomers of KasI. KasI cleaves G\/GCGCC, NarI cuts GG\/CGCC and SfoI cuts to leave blunt GGC\/GCC ends. Both KasI and NarI have site preference, cleaving some sites very slowly. SfoI does not show site preference. Q: Are slow sites found in common vectors? A: Kas is 25-fold more active on lambda than on pBR3222. The slow sites have not been mapped. 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: For enzymes stored at -80°C, is enzyme activity affected after multiple freeze \/ thaw cycles? A: The following enzymes maintained full activity when tested for 10 freeze-thaw cycles. If you require more than 10 freeze-thaw cycles, we recommend aliquoting the enzyme into smaller volumes and freezing each aliquot at -80C. Cac8I EagI-HF I-SceI KasI NlaIII Please note: FseI maintained full activity for only 5 freeze\/thaw cycles. ","brand":"New England Biolabs","offers":[{"title":"Default Title","offer_id":46835536363689,"sku":"R0544S","price":0.99,"currency_code":"USD","in_stock":true}],"url":"https:\/\/iright.com\/products\/neb-r0544s","provider":"Iright","version":"1.0","type":"link"}